Non-transpeptidase binding arylthioether β-lactams active against Mycobacterium tuberculosis and Moraxella catarrhalis

Article abstract of DOI:10.1016/j.bmc.2014.11.025

The prevalence of drug resistance in both clinical and community settings as a consequence of alterations of biosynthetic pathways, enzymes or cell wall architecture is a persistent threat to human health. We have designed, synthesized, and tested a novel

Full text of DOI:10.1016/j.bmc.2014.11.025

Bioorganic & Medicinal Chemistry 23 (2015) 632–647
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Non-transpeptidase binding arylthioether b-lactams active against
Mycobacterium tuberculosis and Moraxella catarrhalis
Tim N. Beck a, Dina Lloyd a, Rostislav Kuskovsky a, Jeanette Minah a, Kriti Arora c, Balbina J. Plotkin b,
Jacalyn M. Green b, Helena I. Boshoff c, Clifton Barry III c, Jeffrey Deschamps d, Monika I. Konaklieva a,
⇑
a Department of Chemistry, American University, Washington, DC 20016, USA
b Department of Microbiology and Immunology, Midwestern University, Chicago, IL 60515, USA
c Tuberculosis Research Section, LCID, NIAID, NIH 33 North Drive, Bldg 33, Rm 2W20C, Bethesda, MD 20892, USA
d Naval Research Laboratory, Code 6930 4555 Overlook Ave., Washington, DC 20375, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
The prevalence of drug resistance in both clinical and community settings as a consequence of alterations
of biosynthetic pathways, enzymes or cell wall architecture is a persistent threat to human health. We
have designed, synthesized, and tested a novel class of non-transpeptidase, b-lactamase resistant
monocyclic b-lactams that carry an arylthio group at C4. These thioethers exhibit inhibitory and cidal
activity against serine b-lactamase producing Mycobacterium tuberculosis wild type strain (Mtb) and
multiple (n = 8) b-lactamase producing Moraxella catarrhalis clinical isolates.
Received 2 September 2014
Revised 8 November 2014
Accepted 18 November 2014
Available online 12 December 2014
Keywords:
Ó 2014 Elsevier Ltd. All rights reserved.
Mycobacterium tuberculosis
Moraxella catarrhalis
b-Lactams
Antimicrobial resistance
1. Introduction
wall crosslinking enzymes, N-thiolated lactams are characterized
by a bacteriostatic activity and act through a different mechanism
Traditional b-lactam ring containing antibiotics have as an
essential requirement for activity, that is, the presence of an
ionizable group either in the proximity (carboxylic acid, bicyclic,
penicillin-like structures) or onto (sulfonic acid, monobactams)
the lactam nitrogen of the b-lactam ring.1–3 This is required for
traditional b-lactam compounds to interact with the bacterial
transpeptidases and inhibit peptidoglycan strand cross-linking.4–6
These compounds are losing their efficacy due to the production
of b-lactamases by resistant bacteria.7,8
Until recently, it was generally accepted that for b-lactams to
exert bactericidal activity, they must contain a scaffold, which spe-
cifically has an ionizable group at the lactam nitrogen within 3.6 Å
of the b-lactam carbonyl carbon.4–6 However, there now appear to
be exceptions to this scaffold requirement, since N-alkylthiolated
b-lactams possess inhibitory, although not cidal, antimicrobial
activity and are stable to the hydrolytic activity of b-lactamases.9–13
The range of bacterial genera affected by these N-alkylthiolated
b-lactams is extremely narrow. The afore-mentioned N-alkylthio-
lated b-lactams are highly selective towards Staphylococcus spp.
and Bacillus spp.9–11 However, unlike penicillins, which inhibit cell
of action.13 Reports confirm that once the ionizable group is
removed from the lactam nitrogen, a variety of novel molecular
targets emerge.14–18 While the mechanisms of N-thiolated
b-lactams are still not completely understood, they appear to affect
cellular processes associated with coenzyme
A and lipid
biosynthesis.13
We developed, synthesized and tested a library of beta-lacta-
mase resistant, non-transpeptidase binding monocyclic b-lactams,
some of which have antimicrobial activity against two phylogenet-
ically distant b-lactamase producing bacterial species—Moraxella
catarrhalis (M.cat.) and Mycobacterium tuberculosis (Mtb).19 The
active compounds were substituted at C4 with alkylthio- as well
as arylthio-groups, with amino-, hydroxy- and sulfhydryl-substitu-
ents at the para position on the aromatic ring, and their N-sulfonates
(Fig. 1). The most active compounds were type I azetidinone
thioethers with arylthio group at C4, lacking the sulfonate at the
lactam nitrogen. N-Sulfenation of these lactams did not enhance
their antimicrobial activity (type II, Fig. 1).19 Structure–activity
relationship (SAR) studies of lactams I and II indicate that the
presence of the C4-arylthio group is essential for antimicrobial
activity. In addition, the N1-substituent has an important role.
Taken together, these data demonstrate that lactams with sulfenat-
ed N1, even with a C4-arylthioether, have either comparable or
diminished antibacterial activity against Mtb and M.cat.19
⇑
Corresponding author. Tel.: +1 202 885 1777; fax: +1 202 885 1752.
E-mail address: mkonak@american.edu (M.I. Konaklieva).
http://dx.doi.org/10.1016/j.bmc.2014.11.025
0968-0896/Ó 2014 Elsevier Ltd. All rights reserved.

T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
633
R
R
R
either unsubstituted at N1 (I, Fig. 1) or sulfenated at N1 (II,
Fig. 1). Antimicrobial activities of the compounds unsubstituted
at the N1 lactam were either similar, or increased, when compared
to the activities of compounds sulfenated at N1. Treatment of
b-lactam 2 with benzylisocyanate produced 30. b-Lactam 30 was
significantly more active against both M.cat. and Mtb (MICs of
S
S
S
NH
N
N
H
N
O
O
Ar
O
S(O)3
I
II
III
O
12.5 lg/ml and 6.25 lg/ml, respectively). The addition of a benzyl-
Figure 1. C4-arylthiolactams with antibacterial activity against Mtb and M.cat.
carbamyl group to the lactam nitrogen also resulted in an increase
in activity for M.cat. and Mtb (>16- and 4-fold, respectively; lac-
tams 2 vs 30; Table 1).
The aims of the present study were to: (1) determine the anti-
microbial activity of compounds in which the sulfur atom at C4
was replaced with an alternative heteroatom; (2) determine the
effects of electron-withdrawing groups (EWGs) or an electron-
donating groups (EDGs) at the arylthio—substituent at C4 on anti-
microbial activity; and (3) identify the optimal functional group at
the lactam nitrogen for maximal antimicrobial activity.
To determine the optimal substituents at C4 and N1, we sys-
tematically synthesized and determined the antimicrobial activi-
ties of compounds containing a H versus a carbamyl moiety at
N1 (Table 1). In addition, b-lactam ethers having arylthio-groups
at C4 with both EWGs and EDGs at o-, m- and p-positions were
synthesized then tested. b-Lactams 6, 8–13 and 27 demonstrated
antimicrobial
activity
against
M.cat.
(MIC 6 12.5 lg/ml;
MBC 6 50 g/ml). The addition of
l
a
carbamyl group at N1
2. Chemistry
improved the anti-M.cat. activity (lactams 30–33, 35, 42–44,
Table 1). Moreover, these lactams exhibit targeted narrow spec-
trum activity against M.cat. and Mtb; they were inactive against
b-lactamase-negative Gram positive and Gram negative quality
control strains and b-lactamase-positive clinical isolates (Table 5;
data not shown).
The overall procedures for generating the various C4-substi-
tuted b-lactams are summarized in Scheme 1. The requirement
for an arylthio-group at the C4 was determined by the synthesis
and subsequent antimicrobial testing of compounds having phen-
oxy- and selenophenoxy-substituents at C4. These compounds
were either unsubstituted or carbamylated at N1.
When considering the effect of the XAr1 group on anti-M.cat.
and anti-Mtb activity, the most active compounds had halogenated
aryl moieties on the thioether (X = S) groups at C4 (Scheme 1). For
example, lactams with arylthioether groups at C4 with multiple
fluoro- or chloro- groups exhibited the highest levels of activity,
regardless of whether the substituent on the lactam nitrogen was
a hydrogen or a carbamyl group (Table 1; compounds 6 & 34; 7
& 35, 8 & 36, 9 & 37, 10 & 38, 11 & 39). Interestingly, positioning
of the fluorine in di-fluorinated compounds affects antimicrobial
activity. Lactam 6 (ortho and para) exhibits a 16- and 8- fold higher
level of cidal activity against M.cat. and Mtb, respectively, when
compared to compound 7 (meta and para). We hypothesize that
placement of fluorine atoms at the ortho- and para-positions,
relative to the sulfur atom in the thiophenoxy group at C4, allows
for a positive electrostatic potential at these regions of the mole-
cule, which could be necessary for formation of a halogen bond
to an appropriate base at the binding site. A halogen bond is a non-
covalent bond between a halogen atom (X) and a Lewis base (B),
similar to the H-bond. This type of bond is attributed to the aniso-
tropic distribution of the charge density on the halogen atom,
The effect on antimicrobial activity of various EWGs or EDGs at
different positions on the aromatic ring was determined. This set of
compounds, C4-arylthio-b-lactams 2–27 (Scheme 1, Table 1) was
prepared from 4-acetoxy-2-azetidinone 1 and the corresponding
arylthiols, by a procedure described by Clauss et al.20 and Wasser-
man et al.21 Carbamylation of the lactam nitrogen (lactams 30–70)
with a variety of isocyanates was accomplished by adopting a pro-
cedure from Mulchande et al.23 or by a microwave-based method
developed in our laboratory. b-Lactams 37–39, 55, 56, 59–63,
65–68, 70 were prepared in methylene chloride (5 ml) with
1.2 M equivalents of triethylamine using a CEM microwave appa-
ratus (300 W, 30 °C for 10–45 min). Incubation times varied
depending on the nature of isocyanate and the unsubstituted
lactams. Lactams 28 and 29 were purchased from Sigma–Aldrich
(St. Louis, MO). Lactam 58, with a terminal triple bond, was pre-
pared by the addition of 5-hexanoic acyl chloride to lactam 2.14
3. Results and discussion
resulting in the formation of a positive cap (called the
r-hole) cen-
We recently described the synthesis and characterization of a
family of C4-arylthio-b-lactams having antimicrobial activity
against Mtb and M.cat.19 The initial library of compounds was
tered on the R–X axis. For a given R, the -hole potential on the
r
halogen X in RX becomes more positive in the order F << Cl < Br < I
(following the trend in the halogens’ polarizabilities).24 The
r-hole
XAr1
OAc
a.
XAr1
H
HXAr1, NaHCO3
acetone/water, RT
b. Ar2NCO or RNCO, Et3N
CH2Cl2
ClCO(CH2)3C
H
N
N
N
N
O
H
O
O
R
c.
CH,
1
2 - 29
O
Et3N, CH2Cl2
30 - 57; 59 - 63; 65 - 70 R = (CH2)nAr2
n = 0 or 1
XAr1
X = O, S, Se
Ar1 = PhY, naphtyl, thiophenyl
Y = H, F, Cl, NO2, CF3, SH, SCH3, OCH3
Ar2 = Ph, C6H5F,
C6H5NO2, C6H5OCH3,
naphtyl
N
O
(CH2)3
O
58
64 R =CH2CH3
X =S, Ar1 = Ph
Scheme 1. Synthesis of N-substituted C4 arylthio-, aryloxy- and arylseleno-b-lactams. (a) b-Lactams 2–29 were prepared from the commercially available b-lactam 1 in the
presence of NaHCO3 in acetone/water.20,21 Preparation of lactams 220, 1722 and 2219 have been reported earlier. (b) b-Lactams 30–70, excluding lactam 58 (R2 = CO(CH2)3CCH)
were prepared from b-lactams 2–29 using aryl isocyanates and Et3N in methylene chloride at rt23, or by using a CEM microwave apparatus at 300 W, 30 °C for 20–45 min. (c)
Lactam 58 can be prepared from lactam 2 as described earlier.14

634
T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
Table 1
Antimicrobial activity of C4-arylthio-, C4-arylseleno- and C4-aryloxy-, unsubstituted at N1, and N1-carbamylated b-lactams against Mtb and M.cat. (n = 6)
R1
N
O
R2
R1
#
R2
H
M.cat.a MIC/MBC (
lg/ml)
Mtb MIC (
lg/ml)
#
R2
M.cat. MIC/MBC (
l
g/ml)
Mtb MIC (
lg/ml)
O
C
S
H
N
2
>200
25
30
31
32
33
34
35
12.5/12.5
6.25
F
O
C
H
N
3
4
5
6
7
H
H
H
H
H
200
25
25/50
3.13
NTc
S
F
O
C
H
N
>200
100
>100
3.13
50
25/100
25/25
S
S
O
C
F
H
N
>200
6.25
100
6.25
F
O
C
H
N
1.5/3.13
50/50
1.63/1.63
12.5/25
S
F
F
O
C
H
N
S
S
F
F
F
F
O
C
F
H
N
8
H
1.5/1.5
>100
36
3.1/6.25
>100
F
Cl
O
C
H
N
9
10
11
12
13
14
15
H
H
H
H
H
H
H
12.5/50
12.5/25
6.25/6.25
6.25/12.5
1.5/1.5
25
37
38
39
40
41
42
43
12.5/25
25–50
25
S
F
F
Cl
O
C
H
N
25
6.25/6.25
3.13/6.25
3.125/25
25/25
S
Cl
O
C
H
N
25
25
S
Cl
O
C
S
NO2
H
N
25
25/
125
25
F3C
S
O
C
H
N
P100
6.25
12.5
O
C
CF3
H
N
200
3.125/6.25
1.625/12.5
S
S
O
C
CF3
H
N
100/100
12.5
CF3
CF3
O
C
H
N
S
16
H
12.5/100
>100
44
6.25/6.25
6.25
H3CO
S
O
C
H
N
17
18
H
H
100/>200
100/>200
250
45
46
200/200
200/200
125–250
100
OCH3
O
C
H
N
>100
S

T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
635
Table 1 (continued)
R1
#
R2
H
M.cat.a MIC/MBC (
lg/ml)
Mtb MIC (
lg/ml)
#
R2
M.cat. MIC/MBC (
lg/ml)
Mtb MIC (
125
lg/ml)
O
C
S
OCH3
H
N
19
200/>200
100
47
200/200
OCH3
O
C
H3CO
S
H
N
20
21
22
23
24
25
26
27
28
29
H
H
H
H
H
H
H
H
H
H
100/>200
200/200
25/100
100/100
100/>200
50/50
250
250
6.25/
>100
>250
25
48
49
50
51
52
53
54
55
56
57
100/200
200/>200
50/100
NT
50
OCH3
O
C
H
N
250
6.25/
>100
>100
250
125
25
S
S
OCH3
O
C
S
H
N
O
C
SMe
S
H
N
O
C
H
N
200/>200
6.25/6.25
50/>200
50/50
S
O
C
O
NO2
H
N
O
C
O
CF3
H
N
200/200
6.25/6.25
NT
250
25
O
C
Se
H
N
O
C
H
N
H
NT
>100
>100
O
C
S
H
N
>200
>200
NT/chemically unstable
NT/chemically unstable
b All compounds are tested as racemates.
a
The values for M.cat. are the results from the six isolates tested. The MIC/MBCs reported were within 2-fold dilutions for all isolates tested regardless of compound. The
concentrations reported are the highest MIC/MBC measured for that compound.
c
NT = not tested.
potential is also more positive as R is more electron-withdrawing.
The more positive the potential -hole of X becomes, the more the
relative to its counterpart lacking the carbamyl-group at N1, and/
or allowed for somewhat increased electrostatic potential on the
halogen atom of the aromatic ring.
r
binding energy of the complex R–X. . .B increases for a given B.24
The probability that the electrostatic potential of the halogens at
the aromatic ring of our compounds may play a role for their anti-
microbial activity is supported by the findings that the di-fluori-
nated lactams 6 and 7 have better antimicrobial activity than
their mono-fluorinated analogs (lactams 3–5). In contrast, the fluo-
rine at the meta position appears to have decreased capability to
form an appropriate halogen bond, based on its location in the aro-
matic ring. This might explain observed differences in antimicro-
bial activity (Table 1, compounds 6 and 7).
To further explore the mechanism by which halogens affect
antimicrobial activity, lactams with chlorine-substituted thiophe-
nols were synthesized and tested for antimicrobial activity. When
one or both fluorines in the thiophenoxy group at C4 were replaced
by chlorine atoms, the lactams demonstrated antimicrobial activ-
ity, regardless of the R2 group (Table 1, compounds with hydrogen
9–11 vs compounds with carbamyl groups 37–39). However,
substitution of a chlorine for a fluorine, in general, decreased
antimicrobial activity of the di-halogenated compounds. Of all
di-halogenated compounds tested compound 6 demonstrated the
most potent antimicrobial activity. The small but measurable
difference in activity between lactams 7 and 10 may be due to
the higher positive electrostatic potential of Cl as compared to F.
The effects of CF3 at various positions relative to the S atom
indicate the differences between M.cat. and Mtb in sensitivity to
these compounds (Table 1). When considering the antimicrobial
activity of compounds 13–16, lactam 13 demonstrated the best
Monofluorinated thiophenoxy groups at C4 alone were insuffi-
cient to make the compounds active within the accepted therapeu-
tic range for screening of novel antimicrobials (MBC <35 lg/ml;
Table 1, compounds 3–5).25,26 However, the replacement of the
proton with a carbamyl group at N1 resulted in significantly
improved antimicrobial activity (Table 1, compounds 31–33 vs
3–5). This improvement in antimicrobial activity could be
explained by several different mechanisms. It may be that the
carbamyl group ‘regenerated’ the isocyanate within the given
microorganism (Scheme 3, mechanism b). Alternatively, it is
plausible that the weak electron-withdrawing effect of the benzyl
carbamyl group increased the electrophilicity of the lactam ring,
activity against M.cat. (MIC 1.5
lg/ml), while lactams 14 and 15
were more effective against Mtb (MIC 6.25–12.5
l
g/ml). When
comparing lactams 4 and 14, the presence of a CF3 group at the
meta-position increased the anti-Mtb activity. This was likely due

636
T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
Table 2
Antimicrobial activity of N1-carbamylated b-lactams against Mtb and M.cat. (n = 6)
R1
N
S
S
F
O
R2
R1=
(2,30,58-61) R1=
(5,62,63)
#
R2
H
M.cat. MIC/MBC (
lg/ml)
Mtb MIC (
lg/ml)
2
>200
25
O
C
H
30
58
12.5/12.5
100/100
6.25
45
N
O
C
O
H
N
C
59
60
NTc
50
O
C
H
N
NT
>100
O
C
H
N
61
5
NT
25
H
>200
>100
O
C
H
N
62
63
>200/>200
>200/>200
>100
>100
O
C
H
N
a The values for M.cat are the results from the six isolates tested. The MIC/MBCs reported were within 2-fold dilutions for all isolates tested
regardless of compound. The concentrations reported are the highest MIC/MBC measured for that compound.
b All compounds are tested as racemates.
c
NT = not tested.
to the higher electron-withdrawing power of the CF3. Hence, the
induction of the CF3 group may decrease even further the electron
density of the atom (in this case S) at C4. The presence of one ver-
sus two CF3 groups at the meta-positions affects the antimicrobial
activity against M.cat. and Mtb but in opposing directions (Table 1,
lactam 14 vs 16). Interestingly, carbamylated 14 and 16, that is,
lactams 42 and 44 (Table 1), demonstrated potent antimicrobial
carbamylation (compound 24 vs 52). This lack of activity may be
due to steric limitations for the bulkier thionaphthyl group, or to
its weak electron-withdrawing capacity. Compound 23 lacked
antimicrobial activity as well, even after carbamylation (51). Inter-
estingly, compound 22, with a 2-thiophenethio-moiety at C4, had
modest (M.cat.) to good (Mtb) antimicrobial activity which was rel-
atively unaffected by carbamylation (lactam 50).
activity against M.cat. and Mtb (MIC 3.13–6.25
25 g/ml, respectively). Substitution of another EWG, para-nitro
lactam, at the thioether moiety at C4, had activity within therapeu-
tic range (<35
g/ml)25,26 for both Mtb and M.cat., regardless of the
presence or absence of a carbamyl group at N1 (Table 1, com-
pounds 12 and 40).
Similar in size to the CF3 group the methoxy group, in contrast,
is an electron donor. The antimicrobial effect of one or more meth-
oxy groups at all positions on the thioaryl moiety at C4 was deter-
mined (Table 1, compounds 17–21 and 45–49). However, this
group of compounds lacked significant activity, even after carba-
mylation at N1 (compounds 45–49). In addition, a lactam with a
thionaphthyl group at C4 lacked antimicrobial activity, even after
l
g/ml and 6.25–
The antimicrobial activities of lactams with aryl groups substi-
tuted at C4, and linked to the lactam ring via an O or Se, were
assessed (Scheme 1; Table 1, compounds 25–27, 53–55). With the
exception of lactams 27 and 53, none of these aryloxy lactams, even
after carbamylation, possessed improved antimicrobial activity rel-
ative to their C4 arylthiol counterparts (Table 1, compounds 25 vs
12; 26 vs 15; 54 vs 43; 55 vs 30). It is likely that the absence of a sul-
fur or selenium atom at the phenyl substituent at C4 accounts for the
lack of activity. Compound 53 has a nitro-substituted phenoxide at
C4, while compound 27 has an arylseleno group at C4, and is unsub-
stituted at N1. Compared to oxygen, sulfur and seleniums are larger
and have greater polarizability. This may explain why sulfur and
selenium, in thiophenoxy and selenophenoxy—substituted at C4
l
l

T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
637
lactams, respectively, carry a larger share of the charge in the transi-
a)
b)
Lactam and thiol products
S
tion state, than oxygen carries in analogous oxygen isosters.27 Thus,
this size/polarizability differential appears to improve antimicrobial
activity. This hypothesis is supported by the charges at X and C4 and
bond lengths of C4-X (Table 4, X = O, S). When compared to the bond
lengths and angles of all non-cyclic thioethers in the Cambridge
Structural Database, the bond lengths and angles of the C4-thio-
phenyl b-lactams described here showed good agreement with the
rest of the known non-cyclic thioethers (Table 4). A similar analysis
showed that ethers and thioethers have bond angles of 116.71° and
102.37°, respectively.
N
H
N
b)
O
Base
a)
Lactam and thiol products
+
Nu
O
Nu =ROH
O
C
N
Scheme 3. Probable reaction pathways of the carbamylated at N1 arylthio-lactams.
In addition to mechanisms (a) described in Scheme 2; mechanism (b) leads to
generation of isocyanate.
Diminished antibacterial activity resulted from replacement of
the N-carbamyl moiety with a carbonyl group (Table 2, lactam 30
vs 58). The intrinsically high chemical reactivity of the lactam ring
in this structure due to the presence of the carbonyl group at N1
may explain these findings. Consistent with this rationale, purifica-
tion of lactam 58 proved difficult due to instability of the lactam ring.
The majority of the lactams having phenoxy-substituents at C4
lacked antimicrobial activity at therapeutic levels. However, the
selenoxy-substituted lactams were bactericidal, even when lacking
an EWG on the aromatic ring (Table 1, 27 and 55). This suggests
that the transition states for loss of thiophenoxide and especially
for selenophenoxide ions from the thiophenoxy- and selenophen-
oxy-substituted at C4 lactams most likely occured later than those
of comparable phenoxy derivatives. This may be a factor determin-
ing the antimicrobial activity of these compounds (Scheme 1).
The findings herein support two possible mechanisms for reac-
tion of nucleophiles with arylthio lactams, unsubstituted at N1
(Scheme 2). In the first possible mechanism, an enzyme having a
serine as a nucleophile hydrolyses the lactam ring; this is a well
documented process (Scheme 2, mechanism a).28,29 Alternatively,
the thiophenol may be the microbicidal part of the molecule with
the lactam moiety acting as the thiophenol carrier (Scheme 2,
mechanism b). Due to the unsubstituted lactam nitrogen the first
mechanism is unlikely to involve the active site serine of the trans-
peptidases, since these lactams lack the necessary prerequisites for
binding to transpeptidases or b-lactamases, that is, a chargeable
polar group at the lactam nitrogen.1–6 Currently, only one monocy-
clic b-lactam having a phenylthioether at C4 and carbamylated at
N1 has been reported to have activity against a pathogen (human
cytomegalovirus).30 As expected, none of the compounds herein
were hydrolysed by penicillinase (data not shown). However, our
compounds were hydrolyzed by sodium hydroxide with departure
of the thiophenoxide from the lactam.27
electron deficiency on sulfur prior to departure, the EWG will cause
the thioether at C4 to act as a leaving group upon deprotonation of
the lactam hydrogen (Scheme 2, mechanism a). In the case of the
lactam’s ‘classical’ enzymatic hydrolysis (Scheme 2 mechanism
b), the presence of the electron withdrawing group(s) will
accelerate these reactions. When N1 is carbamylated mechanism
b in Scheme 3 should also be considered in addition to the
mechanisms described earlier (Scheme 2, mechanism a). As shown
in Scheme 3, an isocyanate molecule could also be released, which
would generate an electrophile; this in turn can bind to a second
nucleophile (Scheme 2, similarly to mechanism b).
Compounds with various R2 substituents at the lactam N1 were
screened for antimicrobial activity to test the mechanism of action
and determine which of our hypotheses were valid. The presence of
a benzyl carbamyl group as the R2 group increased the antimicro-
bial activity. This increase was most likely because carbamylation
makes the lactam ring more electrophilic (Table 1, 2 vs 30; 3–5 vs
31–33; 15–16 vs 43–44). However, the cumulative effect of the
EWG at C4 and the benzyl carbonyl group at N1 did not always cor-
relate with increased antimicrobial activity (Table 1, 6 vs 34; 8–11
vs 36–39; 27 vs 55). Data indicate that if the electron-withdrawing
ability of the substituent at C4 is sufficient for therapeutic levels of
activity, carbamylation at N1, in general, did not increase activity. A
comparable pattern occured when an EWG was attached to the
arylthio moiety of the benzylcarbamyl group. However, if the sub-
stituent at C4 lacked sufficient electron-withdrawing capacity for
the compound to be active, then the substituent at N1 contributed
to the activity. This change in the antimicrobial activity is most
likely the result of increased electrophilicity of the lactam ring.
Compounds with alternate EWGs at N1 and minimal electron
withdrawing capacity at C4 were subsequently synthesized and
tested for antimicrobial activity. M.cat. and Mtb responded
Regardless of the mechanism, the presence of a EWG would be
expected to increase the reactivity of the lactam. By increasing the
Nu2
G
X
-
G
X
G
a)
+
N
Nu1 H
N
Nu1
X
O
H
O
G
3
2
4
b)
1
N
O
H
Base
a)
Nu
Nu1
b)
-
X
Nu1 = ROH
BaseH
+
+
X = O, S, Se
G = H, EWG, EDG
N
O
Scheme 2. Possible reaction pathways of nucleophiles with the arylthio lactams, unsubstituted at N1. (a) ‘Classical’ b-lactam nucleophilic attack on the lactam carbonyl
carbon; phenoxide, (X = O), or thiophenoxide (X = S) or selenophenoxide (X = Se) as a leaving group. (b) Deprotonation of the lactam nitrogen by a base, with departure of a
suitable leaving group, that is, a phenoxide, thiophenoxide or selenophenoxide, followed by protonation to phenol, thiol, or selenol, respectively. The formation of an imine
via pathway (a) as well as the formation of an acyliminium species, a possible Michael acceptor, in (b) allows for a ‘double hit’ mechanism by a second Nu, such as the active
site cysteine in the Mtb glutaminase.

638
T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
Table 3
Antimicrobial activity of fluorinated C4-arylthio-N1-carbamylated b-lactams against Mtb and M.cat. (n = 6)
R1
F
Cl
N
S
F
S
F
O
R2
R1=
(6,34,64-69) R1=
(9, 70)
#
R2
H
M.cat. MIC/MBC (
lg/ml)
Mtb MIC (lg/ml)
6
1.5/3.13
3.13
O
C
H
34
64
1.63/1.63
NTc
100
N
O
>100
>100
C
O
NH
H
N
C
65
66
>200/>200
O
C
H
N
NT
50
F
O
C
H
N
67
68
NT
NT
P100
H CO
3
O
C
H
N
25
F
O
C
H
N
69
9
NT chemically unstable
12.5/50
NT chemically unstable
25
O N
2
H
O
C
H
N
70
>200/>200
50
a The values for M.cat. are the results from the six isolates tested. The MIC/MBCs reported were within 2-fold dilutions for all isolates tested
regardless of compound. The concentrations reported are the highest MIC/MBC measured for that compound.
b All compounds are tested as racemates.
c
NT = not tested.
with phenyl- or naphthyl- and even larger carbamyl groups at
Table 4
Comparison of bond lengths and bond angles of b-lactams with C4 thioethers
described here, with known non-cyclic thioethers and ethers
the N1 (Table 2, 30 vs 58–63). Several of these lactams were
chemically unstable (Table 2, compounds 58; Table 3, 68 and
69); therefore, the optimal substitution remains the benzyl
carbamyl group (Table 1, 30 and 34). Placing an EDG on the
arylthio moiety of the benzyl carbamyl group at N1 does not alter
the activity of the compound, as compared to its benzylcarbamylated
counterpart (Table 3, 67 vs 34).
Compound
S–C(lac) bond
lengths
S–C(R1) bond
lengths
C–S–C bond
angles
2
22
1.803
1.816
1.819
1.819
1.789
1.380
1.773
1.748
1.750
1.750
1.777
1.418
102.37
102.24
102.65
102.64
102.37
116.71
Thio-ethersa
Ethersb
4. Summary
a
Average of all thio-ethers in CSD which do not contain
(n = 7154) from the Cambridge Structural Database.
a
cyclic S-atom
Herein we describe the synthesis and characterization of
C4-arylthio-b-lactams carbamylated at N1, a new family of anti-
bacterial agents with specific activity against b-lactamase produc-
ing Mtb and M.cat. While the presence of a carbamyl substituent at
N1 enhances activity, its presence is not necessary if there are
EWGs on the arylthioether moiety at C4, or halogen atoms with
sufficient electropositive potential for formation of a halogen bond
b
Average of all ethers in CSD which do not contain a cyclic O-atom (n >65,000)
from the Cambridge Structural Database.
differently to compounds with EWGs on the N1-carbamyl. In gen-
eral, the introduction of a benzyl carbamyl group at N1 of the lac-
tam demonstrated better antimicrobial activity than compounds

T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
639
with a base within the active site. The incorporation of EWGs ver-
sus EDGs in the aromatic ring of the arylthio group at C4 led to the
observation that F, Cl, and CF3 substituents at ortho and/or para
positions, in addition to di-substituted compounds at these posi-
tions, possess antibacterial activity within therapeutic range for
novel compounds. The replacement of the urea moiety at N1 with
a carbonyl group, or replacement of the aromatic ring at C4 with an
allyl group, demonstrated diminished antibacterial activity. Based
on our mechanistic investigations, and the lack of prerequisite
groups for binding to the transpeptidase enzymes, these novel
lactams likely have a different mechanism of action from the
b-lactams in current use. Studies are ongoing to optimize the
design and synthesis of the present series of C4-arylthio-, N1-carb-
amylated b-lactams.
GA), preparatory chromatography plates (silica G prep TLC plates
m thickness, Sorbent
Technologies, Inc., Atlanta, GA) and/or recrystallization.
with UV254, 20 Â 20 nm glass backed, 1000
l
All compounds were characterized by 1H and 13C NMR spectra
(25 °C). Spectra were obtained at 400 MHz for 1H NMR and
125 MHz for 13C NMR in CDCl3 or acetone-d6 (Bruker 400 spec-
trometer, Billerica, MA). Chemical shifts were reported in ppm (d)
relative to the residual solvent peak in the corresponding spectra;
chloroform d 7.26 and d 77.23 and acetone d 2.05 coupling
constants (J) are reported in hertz (Hz). Abbreviations used follow:
s = singlet, br = broad singlet, d = doublet, dd = double of doublet,
bd = broad doublet, ddd = doublet of doublet of doublet, t = triplet,
dt = doublet of triplet, q = quartet, p = pentet, m = multiplet. In
most cases, signals due to exchangeable protons were omitted.
IR spectra were obtained from thin films (NaCl plates) and from
solid samples (KBr standard) using
(Columbia, MD). Elemental analyses were performed by Atlantic
Microlab, Inc. (Norcross, GA).
a Shimadzu FT-IR-8300
5. Experimental
5.1. General chemistry methods
Melting points were determined using TA Instruments Q-2000
Differential Scanning Calorimeter (New Castle, DE connected to a
cooling system). Samples (3.0 0.2 mg) were heated or cooled at
a scan rate of 10 °C/min while purged with 50 ml/min N2. For com-
pounds that did not recrystallize, temperatures during the first
heating scan were reported.
Unless stated otherwise, solutions in organic solvents were
dried with anhydrous magnesium sulfate, 37 °C and concentrated
under vacuum conditions using rotator evaporation. Abbrevia-
All air- or moisture-sensitive reactions were performed under
argon or nitrogen atmosphere. Irradiation reactions were
performed in a Discover LabMate from CEM (Weddington, NC).
Reactions were monitored by thin-layer chromatography (TLC)
using EM Reagent plates with fluorescence indicator (SiO2-60,
F
254; thickness 250 lm; EMD Chemicals, Inc., Gibbstown, NJ).
Unless otherwise noted, the compounds were detected under UV
light (254 nm) and iodine vapors.
General methods of compound purification involved flash
chromatography by gradient elution from silica gel columns
(60 Å, particle size 40–75 lm, Sorbent Technologies, Inc., Atlanta,
tions:
DCM = dichloromethane;
DMF = dimethylformamide;
ACN = acetonitrile; EtOAc = ethyl acetate. All compounds were
>98% pure by elemental analysis. Anhydrous solvents, reagent
grade solvents for chromatography and starting materials were
obtained from various sources (Sigma–Aldrich, St. Louis, MO;
Fisher Scientific, Pittsburgh, PA; Aldrich Chemical Co., Milwaulkee,
WI; Matrix Scientific, Columbia, SC; Acros Organics, Geel, Belgium).
Table 5
Organisms tested
Organism
Cell wall type
Acid fast
b-Lactamase
production
5.1.1. General procedure for synthesis of b-lactams containing
aromatic thiols, phenol and benzeneselenol
Mycobacterium tuberculosis H37Rv
BlaC
Moraxella catarrhalis 6 clinical isolates Gram negative
BRO-1/BRO-2
None
None
None
None
Broad spectrum
b-lactamases
Broad spectrum
b-lactamases
ESBLa
ESBL
AmpC
b-lactamase
AmpC
b-lactamase
AmpC
b-lactamase
Metallo
b-lactamases
ESBL &
Carbapenemase
ESBL &
Carbapenemase
ESBL &
Synthesis of compounds 2–28 was performed using procedures
adapted from Grimm and co-workers20 and Wasserman et al.21 To
a solution of 4-acetoxy-2-azetidinone 1 (1 g; 8 mmol in 50 ml ace-
tone/water, 3:2) 1.05 mol equiv of thiophenols, phenol or benzene-
selenol, respectively, were added. Sodium bicarbonate (4 mol equiv)
was added and the mixture was stirred vigorously (12 h; closed
round bottom flask). Sodium chloride was added to the solution
until two layers were formed. The mixture was then filtered and
extracted (EtOAc; 3 Â 50 ml). The combined organic layers were
dried over MgSO4 and concentrated under vacuum. The crude
product was purified by a variety of methods (preparative HPLC;
flash chromatography; and/or recrystallization).
Staphylococcus aureus ATCC 25923
Escherichia coli ATCC 25922
Enterococcus faecalis ATCC 29212
Pseudomonas aeruginosa ATCC 27853
E. coli MISC 206
Gram positive
Gram negative
Gram positive
Gram negative
Gram negative
E. coli GB 8
Gram negative
E. coli MISC 262
Citrobacter freundii Citro 314
Klebsiella pneumoniae MISC 304
Gram negative
Gram negative
Gram negative
K. pneumoniae KLEB 249
C. freundii CITRO 21
Gram negative
Gram negative
Gram negative
Gram negative
Gram negative
Gram negative
Gram negative
Gram negative
Gram negative
Gram negative
5.1.1.1. 4-Phenylsulfanyl-azetidin-2-one (2).
The crude
Stenotrophomonas maltophilia GM24
Acinetobacter baumannii L185
Acinetobacter baumannii L186
Acinetobacter baumannii L187
K. pneumoniae/KPC L133
K. pneumoniae/ESBL L174
E. coli ESBL L108
product was crystallized from CH2Cl2 and hexanes to afford white
crystals (62%) with mp 66–68 °C. 1H NMR (400 MHz, CDCl3): d 7.42
(5H, m), 6.1 (1H, br s), 5.02 (1H, dd, J = 2.3, 4.9), 3.38 (1H, ddd,
J = 1.9, 4.9, 15.2), 2.9 (1H, ddd, J = 1.5, 2.3, 15.2); 13C NMR
(100 MHz, CDCl3) d 165.56, 133.60, 131.21, 129.43, 128.76, 54.20,
45.40. IR (neat) m
max (C@O): 1766.7 cmÀ1. Anal. Calcd for C9H9NOS:
Carbapenemase
ESBL &
Carbapenemase
ESBL &
Carbapenemase
ESBL &
Carbapenemase
ESBL &
C, 60.31; H, 5.06; N, 7.81. Found: C, 60.26; H, 5.04; N, 7.81.
5.1.1.2. 4-(2-Fluoro-phenylsulfanyl)-azetidin-2-one (3).
The
product was purified by flash chromatography from ethyl ace-
tate/hexanes to give white crystals (65%) yield with mp
101.83 °C. 1H NMR (400 MHz, CDCl3): d 7.30 (5H, m), 6.16 (1H, br
s), 4.99 (1H, dd, J = 2.3, 5.0), 3.36 (1H, ddd, J = 2.4, 7.8, 15.7), 2.87
(1H, dd, J = 1.3, 7.8, 15.7).
E. coli ESBL L109
Carbapenemase
a
ESBL = extended spectrum b-lactamase.

640
T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
13C NMR (100 MHz, CDCl3) d 165.75, 164.02, 161.57, 136.30,
5.1.1.6. 4-(3,4-Difluoro-phenylsulfanyl)-azetidin-2-one (7). The
product was purified by recrystallization in ethyl acetate/hexanes to
give white crystals (47.1%) with mp 74–77 °C. 1H NMR (400 MHz,
CDCl3): d 7.11–7.37 (3H, m), 6.17 (1H, br s), 5.00 (1H, dd, J = 2.3,
4.9), 3.64 (1H, ddd, J = 1.7, 7.8, 15.7), 2.90 (1H, ddd, J = 1.6, 7.8,
15.7); 13C NMR (100 MHz, CDCl3) d 166.53, 151.87, 149.89, 130.77,
127.39, 123.23, 118.33, 54.70, 45.45; IR (neat) mmax (C@O)
1750 cmÀ1. Anal. Calcd for C9H7F2NOS: C, 50.23; H, 3.28; N, 6.51.
Found: C, 50.15; H, 3.26; N, 6.45.
131.40, 125.06, 116.62, 114.89, 54.44, 46.04; IR (neat) mmax (C@O)
1750 cmÀ1; X-ray analyses.
5.1.1.7. 4-Pentafluorophenylsulfanyl-azetidin-2-one (8). The
product was purified by recrystallization in ethyl acetate/hexanes
to give white crystals (46%) with mp 80–82 °C. 1H NMR (400 MHz,
CDCl3): d 6.75 (1H, s), 4.91 (1H, dd, J = 2.3, 4.9), 3.40 (1H, ddd,
J = 2.4, 7.9, 15.2), 2.92 (1H, ddd, J = 1.6, 7.9, 15.3); 13C NMR
(100 MHz, CDCl3) d 165.81, 150.54, 148.98, 140.59, 136.05, 112.26,
51.63, 46.26; IR (neat) m
max (C@O) 1755 cmÀ1. Anal. Calcd for C9H4F5
NOS: C, 40.16; H, 1.50; N, 5.20. Found: C, 40.36; H, 1.79; N, 4.97.
5.1.1.8. 4-(2-Chloro-4-fluoro-phenylsulfanyl)-azetidin-2-one (9). The
product was purified by recrystallization in ethyl acetate/hexanes
to give white crystals (66%) mp 84–85 °C. 1H NMR (400 MHz,
CDCl3): d 7.56 (1H, dd, J = 0.5, 8.8), 7.20 (ddd, J = 0.5, 2.6, 5.5),
7.04 (1H, ddd, J = 2.6, 5.5, 8.8), 6.46 (1H, br s), 5.02 (1H, dd,
J = 2.4, 4.8), 3.45 (1H, ddd, J = 1.8, 3.0, 8.5), 2.97 (1H, td, J = 1.8,
11.6); 13C NMR (100 MHz, CDCl3): d 165.65, 164.32, 161.78,
137.58, 126.45, 118.63, 115.09, 54.36, 45.81; IR (neat) mmax (C@O)
1756.93 cmÀ1. Anal. Calcd for C9H7ClFNOS: C, 46.66; H, 3.05; Cl,
15.30; F, 8.20; N, 6.05; O, 6.91; S, 13.84. Found: C, 46.97; H, 3.23;
Cl, 15.51; F, 8.13; N, 6.22; O, 6.71; S, 13.64.
5.1.1.3.
4-(3-Fluoro-phenylsulfanyl)-azetidin-2-one
(4).
The product was purified by flash chromatography from ethyl
acetate/hexanes to give white crystals (65%) with mp 60.94 °C. 1H
NMR (400 MHz, CDCl3): d 7.30 (5H, m), 6.16 (1H, br s), 4.99 (1H,
dd, J = 2.3, 4.9), 3.36 (1H, ddd, J = 1.8, 7.8, 15.7), 2.87 (1H, dd,
J = 1.3, 7.8, 15.7); 13C NMR (100 MHz, CDCl3) d 165.95, 160.92,
130.93, 128.60, 119.92, 119.70, 115.67, 54.21, 45.81; IR (neat) mmax
(C@O) 1750 cmÀ1. Anal. Calcd for C17H14F2N2O2S: C, 58.61; H, 4.05;
N, 8.04. Found: C, 58.29; H, 3.99; N, 8.07.
5.1.1.4. 4-(4-Fluoro-phenylsulfanyl)-azetidin-2-one (5). The
product was purified by flash chromatography from ethyl
acetate/hexanes to give white crystals (70%) with mp 76.67 °C.
1H NMR (400 MHz, CDCl3): d 7.26 (2H, ddd, J = 0.5, 2.0, 8.9), 6.99
(2H, ddd, J = 0.5. 5.1, 8.9), 6.11 (1H, br s), 4.96 (1H, dd, J = 2.4,
4.8), 3.37 (1H, ddd, J = 1.8, 7.9, 15.7), 2.87 (1H, ddd, J = 1.6, 7.8,
5.1.1.9. 4-(3-Chloro-4-fluoro-phenylsulfanyl)-azetidin-2-one (10) . The
product was purified by recrystallization in ethyl acetate/hexanes
to give white crystals (56%) with mp 97–98 °C. 1H NMR
(400 MHz, CDCl3): d 7.57 (1H, dd, J = 0.5, 2.5), 7.39 (1H, dd,
J = 2.2, 8.9), 7.20 (1H, ddd, J = 0.5, 5.1, 8.6), 6.28 (1H, br s), 4.99
(1H, dd, J = 2.3, 4.9), 3.42 (1H, ddd, J = 2.3, 7.8, 15.4), 2.90 (1H,
ddd, J = 1.2, 7.8, 15.8); 13C NMR (100 MHz, CDCl3) d 165.54,
159.99, 136.29, 134.31, 130.82, 127.34, 121.99, 117.46, 54.58,
15.2); 13C NMR (100 MHz, CDCl3)
d 166.73, 162.21, 136.63,
126.13, 116.59, 54.78, 45.28; IR (neat) mmax (C@O) 1750 cmÀ1
.
X-ray analysis.
45.47; IR (neat) m
max (C@O) 1761.81 cmÀ1 Anal. Calcd for C9H7ClF-
NOS: C, 46.66; H, 3.05; Cl, 15.30; F, 8.20; N, 6.05; O, 6.91; S, 13.84.
Found: C, 46.57; H, 3.28; Cl, 15.41; F, 8.27; N, 6.28; O, 6.86; S,
13.76.
5.1.1.10. 4-(2,4-Dichloro-phenylsulfanyl)-azetidin-2-one (11).
The product was purified by recrystallization in ethyl acetate/
hexanes to give white crystals (50%) yield with mp 143–144 °C.
1H NMR (400 MHz, CDCl3): d 7.53 (1H, dd, J = 0.5, 2.2), 7.43
(1H, dd, J = 2.2, 8.3), 7.29 (2H, dd, J = 0.5, 8.5), 6.32 (1H, br s),
5.70 (1H, dd, J = 2.3, 4.9), 3.50 (1H, ddd, J = 1.9, 8.5, 15.7), 3.02
(1H, ddd, J = 1.5, 7.9, 15.5); 13C NMR (100 MHz, CDCl3)
d
165.32, 138.25, 135.49, 135.20, 130.35, 129.65, 127.97, 53.90,
46.02; IR (neat) mmax (C@O) 1683.76 cmÀ1. Anal. Calcd for C9H7
Cl2NOS: C, 43.57; H, 2.84; Cl, 28.57; N, 5.65; O, 6.45; S, 12.92.
Found: C, 43.77; H, 2.98; Cl, 28.73; N, 5.57; O, 6.37; S, 12.84.
5.1.1.5. 4-(2,4-Difluoro-phenylsulfanyl)-azetidin-2-one (6). The
product was purified by recrystallization in ethyl acetate/hexanes
to give white crystals (70%) with mp 70–73 °C. 1H NMR
(400 MHz, CDCl3): d 7.52 (1H, dd, J = 0.5, 8.8), 6.85 (1H, ddd,
J = 2.7, 5.1, 8.8), 6.78 (1H, ddd, J = 0.5, 2.7, 5.1), 6.13 (1H, br s),
4.96 (1H, dd, J = 2.3, 4.9), 3.42 (1H, ddd, J = 1.8, 8.5, 15.7), 2.29
(1H, ddd, J = 1.8, 7.8, 15.6); 13C NMR (100 MHz, CDCl3) d 166.43,
165.01, 161.90, 139.36, 112.31, 11.34, 104.88, 54.38, 45.55; IR
(neat) mmax (C@O) 1750 cmÀ1. Anal. Calcd for C9H7F2NOS: C,
50.23; H, 3.28; N, 6.51. Found: C, 50.32; H, 3.30; N, 6.35.
5.1.1.11. 4-(4-Nitro-phenylsulfanyl)-azetidin-2-one (12). The
product was purified by recrystallization in ethyl acetate/hexanes
to give white crystals (81%) with mp 124.03 °C. 1H NMR (400 MHz,
CDCl3): d 8.20 (2H, dd, J = 2.0, 8.4), 7.48 (2H, dd, J = 3.1, 8.5), 6.24
(1H, br s), 5.22 (1H, dd, J = 2.3, 4.9), 3.58 (1H, ddd, J = 1.7, 8.7,
15.7), 3.05 (1H, ddd, J = 1.5, 7.9, 15.7); 13C NMR (100 MHz, CDCl3)
d 165.95, 130.93, 128.60, 119.92, 119.70, 115.88, 115.67, 54.21,
45.81; IR (neat)
C, 48.21; H, 3.60; N, 12.49. Found: C, 48.16; H, 3.50; N, 12.39.
m
max (C@O) 1700 cmÀ1. Anal. Calcd for C9H8N2O3S:

T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
641
5.1.1.12.
(13).
4-((2-(Trifluoromethyl)phenyl)thio)azetidin-2-one
The product was purified by recrystallization in ethyl
5.1.1.18. 4-(4-Methoxy-phenylsulfanyl)-azetidin-2-one (19). The
product was purified by recrystallization in hexanes/ethyl acetate
to give a white solid (57%) with mp 83–85 °C. 1H NMR (400 MHz,
CDCl3): d 7.40–6.86 (4H, m), 6.81 (1H, br s), 4.97 (1H, dd, J = 2.3,
4.9), 3.89 (3H, s), 3.37 (1H, ddd, J = 2.4, 8.8, 15.7), 2.9 (1H, dd,
J = 1.9, 7.9, 15.7); 13C NMR (100 MHz, CDCl3) d 169.69, 159.16,
134.93, 130.47, 121.44, 111.47, 56.06, 54.06, 45.87; IR (neat) mmax
(C@O) 1756 cmÀ1. Anal. Calcd for C10H11NO2S: C, 57.39; H, 5.30;
N, 6.69. Found: C, 57.20; H, 5.22; N, 6.58.
acetate/hexanes to give a yellow-white powder (90%) with mp
81–82 °C. 1H NMR (400 MHz, CDCl3): d 7.75 (1H, d, J = 7.6), 7.62
(1H, dd, J = 1.3, 7.5), 7.56 (1H, dd, J = 1.5, 7.8), 7.47 (1H, d, J = 7.1),
6.91 (1H, s), 5.03 (1H, dd, J = 2.3, 4.9), 3.44 (1H, ddd, J = 1.8, 7.9,
15.7), 2.95 (1H, ddd, J = 1.6, 7.9, 15.7); 13C NMR (100 MHz, CDCl3)
d 166.26, 135.88, 132.63, 131.42, 128.80, 127.47, 55.05, 45.99; IR
(neat) mmax (C@O) 1766 cmÀ1. Anal. Calcd for C10H8F3NOS: C,
48.58; H, 3.26; N, 5.67. Found: C, 48.73; H, 3.39; N, 5.83.
5.1.1.19. 4-(2,5-Dimethoxy-phenylsulfanyl)-azetidin-2-one (20).
The product was purified by recrystallization in hexanes/ethyl ace-
tate to give a white solid (51%) with mp 116–118 °C. 1H NMR
(400 MHz, CDCl3): d 7.28–6.87 (4H, m), 6.46 (1H, br s), 5.01 (1H,
dd, J = 2.3, 4.9), 3.78 (3H, s), 3.41 (1H, ddd, J = 2.4, 8.8, 15.7), 3.86
(3H, s), 2.98 (1H, ddd, J = 2.0, 8.8, 15.7); 13C NMR (100 MHz, CDCl3)
d 166.26, 153.59, 153.19, 120.93, 120.13, 114.78, 112.22, 56.44,
55.83, 53.88, 45.73; IR (neat) mmax (C@O) 1760 cmÀ1. Anal. Calcd
for C11H13NO3S: C, 55.21; H, 5.48; N, 5.85. Found: C, 54.92; H,
5.36; N, 5.81.13.
5.1.1.13. 4-((3-(Trifluoromethyl)phenyl)thio)azetidin-2-one (14).
The product was purified by recrystallization in ethyl acetate/hex-
anes to give white crystals (83%) with mp 73–75 °C. 1H NMR
(400 MHz, CDCl3): d 7.72 (1H, d, J = 1.9), 7.67 (1H, dd, J = 1.8, 9.2),
7.62 (1H, dd, J = 7.7, 7.9), 7.51 (1H, dd, J = 1.9, 7.7), 6.80 (1H, s),
5.07 (1H, dd, J = 2.3, 4.9), 3.45 (1H, ddd, J = 1.9, 8.5, 15.7), 2.92
(1H, ddd, J = 1,6, 7.9, 15.7); 13C NMR (100 MHz, CDCl3): d 166.39,
136.32, 133.41, 131.95 (1C, q, J = 0.33), 130.03, 129.75 (1C, q,
J = 0.03), 125.43 (1C, q, J = 0.03), 54.38, 45.83; IR (neat) mmax
(C@O) 1766 cmÀ1. Anal. Calcd for C10H8F3NOS: C, 48.58; H, 3.26;
N, 5.67. Found: C, 48.65; H, 3.09; N, 5.63.
5.1.1.20. 4-(3,4-Dimethoxy-phenylsulfanyl)-azetidin-2-one (21).
The product was purified by recrystallization in hexanes/ethyl ace-
tate to give a white powder (63%) with mp 79–80 °C. 1H NMR
(400 MHz, CDCl3): d 7.07–6.34 (4H, m), 6.36 (1H, br s), 4.71 (1H,
dd, J = 2.3, 4.9), 3.68 (3H, s), 3.67 (3H, s), 3.10 (1H, ddd, J = 1.9,
8.4, 15.7), 2.64 (1H, ddd, J = 1.6, 8.3, 15.7); 13C NMR (100 MHz,
CDCl3) d 171.20, 166.21, 150.12, 149.15, 128.05, 121.20, 117.74,
111.59, 60.41, 56.02, 55.91, 54.78, 44.89, 21.06, 14.19; IR (neat)
mmax (C@O) 1760 cmÀ1. Anal. Calcd for C11H13NO3S: C, 55.21; H,
5.48; N, 5.85. Found: C, 55.03; H, 5.40; N, 5.78.
5.1.1.14. 4-((4-(Trifluoromethyl)phenyl)thio)azetidin-2-one (15). The
product was purified by recrystallization in ethyl acetate/hexanes
to give white crystals (81%) with mp 95–96.5 °C. 1H NMR
(400 MHz, CDCl3): d 7.61 (2H, doublet, J = 8.2), 7.51 (2H, doublet,
J = 8.1), 7.01 (1H, singlet), 5.11 (1H, dd, J = 2.3, 4.9), 3.47 (1H,
ddd, J = 1.9, 3.0, 8.4), 2.95 (1H, ddd, J = 0.8, 1.4, 11.7); 13C NMR
(100 MHz, CDCl3)
d 166.43, 137.57, 131.75, 130.13 (1C, q,
J = 0.33), 126.34, 123.79, 53.78, 45.91; IR (neat) mmax (C@O)
1764 cmÀ1. Anal. Calcd for C10H8F3NOS: C, 48.58; H, 3.26; N,
5.67. Found: C, 48.83; H, 3.29; N, 5.53.
5.1.1.21. 4-(Thiophen-2-ylsulfanyl)-azetidin-2-one (22). The
crude product was crystallized from hexanes/methylene chloride
to afford white crystals (90%) with mp 57–58 °C. 1H NMR
(400 MHz,CDCl3): d 7.48 (1H, dd, J = 1.2, 5.4), 7.23 (1H, dd, J = 1.2,
3.6), 7.07 (1H, dd, J = 3.6, 5.4), 6.19 (1H, br s), 4.86 (1H, dd,
J = 2.3, 4.9), 3.31 (1H, ddd, J = 1.9, 4.9, 15.3), 2.91 (1H, ddd, J = 1.6,
2.0, 15.2); 13C NMR (100 MHz, CDCl3) d 165.52, 136.75, 131.82,
5.1.1.15. 4-((3,5-Bis(trifluoromethyl)phenyl)thio)azetidin-2-one
(16).
The product was purified by recrystallization in ethyl
acetate/hexanes to give white crystals (79%) with mp 77–79 °C.
1H NMR (400 MHz, CDCl3): d 7.88 (2H, dd, J = 2.0, 8.3), 7.86 (2H,
dd, J = 1.9, 8.3), 6.46 (1H, s), 5.17 (1H, dd, J = 2.3, 4.9), 3.55 (1H,
ddd, J = 2.4, 8.3, 15.7), 3.00 (1H, ddd, J = 1.6, 7.9, 15.7); 13C NMR
128.12, 127.49, 55.38, 44.72; IR (neat) mmax (C@O) 1739.7.1 cmÀ1
.
Anal. Calcd for C7H7NOS2: C, 45.38; H, 3.81; N, 7.56. Found: C,
45.50; H, 3.76; N, 7.43.
(100 MHz, CDCl3)
d 166.04, 135.93, 132.77 (1C, q, J = 0.33),
132.11, 124.30, 122.11, 121.59, 54.22, 46.04; IR (neat) mmax (C@O)
1771 cmÀ1. Anal. Calcd for C11H7F6NOS: C, 41.91; H, 2.24; N,
4.44. Found: C, 41.83; H, 2.08; N, 4.43.
5.1.1.22. 4-(4-Methylsulfanyl-phenylsulfanyl)-azetidin-2-one (23).
The product was recrystallized from hexanes/ethyl acetate to give
white fluffy powder (49%) with mp 60.16 °C. 1H NMR (400 MHz,
CDCl3) dH 7.27 (2H, dd, J = 2.0, 8.1), 7.0 (2H, dd, J = 2.0, 8.1), 6.07
(1H, br s), 4.97 (1H, dd, J = 2.6, 5.3), 3.36 (1H, ddd, J = 2.4, 7.6,
15.7), 2.88 (1H, ddd, J = 1.6, 7.8, 15.5), 2.50 (3H, s); 13C NMR
(100 MHz, CDCl3) d 166.12, 140.67, 134.80, 126.93, 125.89, 54.63,
45.36, 15.52; IR (neat) mmax (C@O) 1700 cmÀ1. Anal. Calcd for
5.1.1.16. 4-(2-Methoxy-phenylsulfanyl)-azetidin-2-one (17). The
product was purified by recrystallization in hexane/ethyl acetate to
give a white solid (87%) with mp 106–107 °C. 1H NMR (400 MHz,
CDCl3): d 7.43–6.88 (4H, m), 6.75 (1H, br s), 4.86 (1H, dd, J = 2.3,
4.9), 3.80 (3H, s), 3.28 (1H, ddd, J = 1.8, 8.5, 15.7), 2.93 (1H, ddd,
J = 1.4, 8.5, 15.8); 13C NMR (100 MHz, CDCl3) d 166.56, 160.74,
136.93, 120.82, 114.96, 55.54, 54.90, 44.99; IR (neat) mmax (C@O)
1760 cmÀ1. Anal. Calcd for C10H11NO2S: C, 57.39; H, 5.30; N, 6.69.
Found: C, 57.37; H, 5.19; N, 6.67.
C10H11NOS2: C, 53.30; H, 4.92; N, 6.22. Found: C, 53.33; H, 4.90;
N, 6.16.
5.1.1.23. 4-(Naphthalen-1-ylsulfanyl)-azetidin-2-one (24). The
product was purified by recrystallization in hexanes/ethyl acetate
to give white crystals (73%) with mp 124.12 °C. 1H NMR
(400 MHz, CDCl3): d 8.00 (3H, m), 7.87–7.55 (4H, m), 6.175 (1H,
br s), 5.14 (1H, dd, J = 2.5, 5.0), 3.45 (1H, ddd, J = 1.81, 7.9, 15.7),
3.00 (1H, ddd, J = 1.7, 7.8, 15.4); 13C NMR (100 MHz, CDCl3): d
165.42, 149.70, 133.63, 132.90, 132.79, 130.16, 129.14, 128.72,
127.81, 127.62, 126.97, 126.94, 54.29, 45.49; IR (neat) mmax
(C@O) 1755 cmÀ1. Anal. Calcd for C13H11NOS: C, 68.10; H, 4.84;
N, 6.11; O, 6.98; S, 13.98. Found: C, 68.35; H, 4.97; N, 6.19; O,
6.79; S, 13.78.
5.1.1.17. 4-(3-Methoxy-phenylsulfanyl)-azetidin-2-one (18). The
product was purified on
a silica gel column (hexanes/ethyl
acetate, 1:1; v/v) to give yellow oil in 75% yield. 1H NMR
(400 MHz, CDCl3): d 7.29–6.87 (4H, m), 6.07 (1H, br s), 5.03
(1H, dd, J = 2.3, 4.9), 3.81 (3H, s), 3.39 (1H, ddd, J = 2.4, 7.9, 15.7),
2.91 (1H, dd, J = 1.4, 7.9, 15.7); 13C NMR (100 MHz, CDCl3) d
169.99, 160.01, 133.17, 130.30, 125.02, 118.24, 114.22, 55.46,
54.22, 45.41; IR (neat) mmax (C@O) 1756 cmÀ1. Anal. Calcd for
C10H11NO2S: C, 57.39; H, 5.30; N, 6.69. Found: C, 57.25; H, 5.34;
N, 6.52.

642
T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
5.1.1.24. 4-(4-Nitro-phenoxy)-azetidin-2-one (25).
Purified
(6H, m), 7.37 (1H, m), 7.13 (3H, m), 6.85 (1H, br s), 5.32 (1H, dd,
J = 2.3, 4.9), 4.53 (1H, dd, J = 1.0, 17.0), 3.43 (1H, dd, J = 7.5, 15.7),
2.85 (1H, dd, J = 7.6, 16.0); 13C NMR (100 MHz, CDCl3) d 165.44,
149.63, 137.82, 135.27, 129.32, 129.30, 128.78, 127.73, 127.69,
127.70, 100.00, 56.69, 43.99, 43.69; IR (neat) mmax (C@O)
1755 cmÀ1. Anal. Calcd for C17H16N2O2S: C, 65.36; H, 5.16; N,
8.97; Found: C, 65.58; H, 5.45; N, 8.89.
by recrystallization in hexanes/ethyl acetate to give a beige powder
(68%) with mp 124–127 °C. 1H NMR (400 MHz, CDCl3): d 8.28 (2H,
dd, J = 2.8, 9.4), 6.97 (2H, dd, J = 3.1, 9.3), 6.56 (1H, br s), 5.81 (1H,
dd, J = 2.3, 4.9), 3.49 (1H, ddd, J = 1.3, 2.0, 15.0), 3.2 (1H, ddd, J = 1.5,
3.9, 15.1); 13C NMR (100 MHz, (CD3)2CO) d 166.14, 162.45, 143.17,
126.74, 116.62, 77.40, 46.69; IR (neat) m
max (C@O) 1775 cmÀ1. Anal.
Calcd for C9H8N2O4: C, 51.93; H, 3.87; N, 13.46. Found: C, 51.64; H,
4.02; N, 13.46.
5.1.3.2. 2-(2-Fluoro-phenylsulfanyl)-4-oxo-azetidine-1-carbox-
ylic acid benzylamide (31).
The product was purified by flash
5.1.1.25. 4-(4-Trifluoromethyl-phenoxy)-azetidin-2-one (26).
The product was purified by recrystallization in hexanes/ethyl ace-
tate to give a white powder (78%) with mp 95–98 °C. 1H NMR
(400 MHz, CDCl3): d 7.54 (2H, dd, J = 1.9, 8.7), 6.87 (2H, dd,
J = 2.5, 8.6), 6.53 (1H, br s), 5.67 (1H, dd, J = 2.3, 5.0), 3.35 (1H,
ddd, J = 1.4, 1.6, 15.0), 3.07 (1H, d, J = 1.6, 3.9, 15.1); 13C NMR
chromatography on silica gel (hexanes/ethyl acetate, 1:1; v/v) to
afford the compound as white crystals (67%) with mp 89.79 °C.
1H NMR (400 MHz, CDCl3): d 7.61–7.30 (3H, m), 7.18–7.12 (5H,
m), 6.99 (1H, ddd, J = 1.4, 5.1, 8.3), 6.77 (1H, br s), 5.35 (1H, dd,
J = 2.3, 4.9), 4.50 (2H, dd, J = 1.0, 17.0), 3.47 (1H, dd, J = 7.5, 16.1);
2.94 (1H, dd, J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3): d 164.46,
162.00, 160.39, 149.64, 137.95, 137.77, 132.04, 128.82, 127.63,
116.49, 116.27, 56.29, 43.85, 40.74; IR (neat) mmax (C@O) 1774,
1709 cmÀ1. Anal. Calcd for C17H14F2N2O2S: C, 61.80; H, 4.85;
N, 8.45. Found: C, 61.95; H, 4.62; N, 8.45.
(100 MHz, CDCl3)
d 166.46, 158.37, 127.36, 125.11, 124.78,
115.43, 76.23, 46.18; IR (neat) mmax (C@O) 1780 cmÀ1. Anal. Calcd
for C10H8F3NO2: C, 51.96; H, 3.49; N, 6.06. Found: C, 51.91; H,
3.41; N, 5.97.
5.1.1.26. 4-Phenylselanyl-azetidin-2-one (27).
The product
was purified on silica gel (hexanes/ethyl acetate 1:1; v/v) to afford
the compound as white crystals (50%) with mp 78–80 °C. 1H NMR
(400 MHz, CDCl3): d 7.35–7.89 (5H, m), 6.33 (1H, br s), 5.14 (1H, dd,
J = 2.3, 4.9), 3.45 (1H, ddd, J = 1.70, 7.8, 14.8), 2.97 (1H, ddd, J = 1.6,
7.8, 15.6); 13C NMR (100 MHz, CDCl3): d 165.91, 135.79, 131.52,
129.52, 129.23, 127.76, 47.29, 46.46; IR (neat) mmax (C@O)
1749.95 cmÀ1. Anal. Calcd for C9H9NOSe: C, 47.80; H, 4.01; N,
6.19; O, 7.07; Se, 34.92. Found: C, 47.92; H, 4.21; N, 6.39; O,
7.27; Se, 34.89
Lactams 28 (azetidin-2-one) and 29 (4-allylsulfanyl-azetidin-2-
one) were purchased from Sigma–Aldrich (28, 29).
5.1.2. General procedure for the synthesis of N-carbamoylazeti-
din-2-one derivatives (30–36, 40–54, 57, 58, 64, 69 from 2 to 29,
respectively)
The synthetic procedure was adapted from Mulchande et al.23
To a solution of appropriate azetidin-2-one 5 (1.7 g, 5.4 mmol)
in DCM (5 ml) was added to 1.2 mol equiv of triethylamine and
1.2 mol equiv of the corresponding isocyanates: benzyl isocyanate,
ethyl isocyanate, 2-nitrophenyl isocyanate. The reaction was stir-
red at room temperature and monitored by TLC. After completion
of the reaction, the solution was evaporated under reduced
pressure.
5.1.3.3. 2-(3-Fluoro-phenylsulfanyl)-4-oxo-azetidine-1-carbox-
ylic acid benzylamide (32).
The product is a clear yellow oil
purified by flash chromatography on silica gel (hexanes/ethyl ace-
tate, 1:1; v/v) in 70% yield. 1H NMR (400 MHz, CDCl3): d 7.40–7.32
(2H, m),7.16–7.10 (5H, m), 7.1 (1H, dd, J = 7.7, 8.3), 6.84 (1H, ddd,
J = 1.8, 5.1, 8.3), 6.78 (1H br s), 5.33 (1H, dd, J = 2.3, 4.9), 4.52
(2H, dd, J = 1.0, 17.0), 3.46 (1H, dd, J = 7.5, 16.0), 2.94 (1H, dd,
J = 7.5, 15.7); 13C NMR (100 MHz, CDCl3)
d 165.44, 156.15,
149.77, 138.04, 132.60, 130.70, 130.13, 128.95, 127.84, 121.35,
121.13, 116.39, 116.18, 56.98, 44.47, 43.82; IR (neat) mmax (C@O)
1774, 1709 cmÀ1. Anal. Calcd for C17H14F2N2O2S: C, 61.80; H,
4.58; N, 8.44. Found: C, 61.77; H, 4.49; N, 8.44.
5.1.3. General procedure for the synthesis of N-carbamoylazeti-
din-2-one derivatives via irradiation (37–39, 55, 56, 59–63,
65–68, 70)
The synthetic procedure developed in our laboratory using a
CEM microwave apparatus was applied to the aforementioned
compounds: to a solution of an appropriate azetidin-2-one 5
(0.1 g, 0.46 mmol) in dichloromethane (4 ml), 2 mol equiv of tri-
ethylamine was added followed by addition of 1.1 mol equiv of
the corresponding isocyanates: benzyl isocyanate, diphenylmethyl
isocyanate, 9H-fluoren-9-yl isocyanate, 1-naphthyl isocyanate, 2-
fluorobenzyl isocyanate, 2-fluorophenyl isocyanate, 2-methoxy-
benzyl isocyanate, diphenylethyl isocyanate, and 1-(1-naph-
thyl)ethyl isocyanate. The reaction was irradiated under-pressure
in the microwave for 10–60 min at 300 W, and 35 °C and moni-
tored by TLC and NMR.
5.1.3.4. 2-(4-Fluoro-phenylsulfanyl)-4-oxo-azetidine-1-carbox-
ylic acid benzylamide (33).
The product was purified by flash
chromatography on silica gel (hexanes/ethyl acetate, 1:1; v/v) to
afford the compound as white crystals (60%) with mp 89.71 °C.
1H NMR (400 MHz, CDCl3): d 7.58–7.30 (5H, m), 7.15–7.10 (3H,
m) 7.01–6.95 (2H, ddd, J = 2.68, 5.2, 8.9), 6.84 (1H, br s), 5.26
(1H, dd, J = 2.3, 4.9), 4.52 (2H, dd, J = 1.0, 17.0), 3.44 (1H, dd,
J = 7.5, 16.0); 2.89 (1H, dd, J = 7.5, 15.6); 13C NMR (100 MHz, CDCl3)
d 165.44, 165.01, 162.52, 149.79, 137.83, 128.96, 127.88, 124.72,
116.77, 116.56, 57.22, 48.36, 44.07; IR (neat) mmax (C@O) 1774,
1709 cmÀ1. Anal. Calcd for C17H14F2N2O2S: C, 61.80; H, 4.58; N,
8.44. Found: C, 61.75; H, 4.49; N, 8.47.
5.1.3.1. 2-Oxo-4-phenylsulfanyl-azetidine-1-carboxylic acid
5.1.3.5. 2-(2,4-Difluoro-phenylsulfanyl)-4-oxo-azetidine-1-car-
benzylamide (30).
The product was purified on silica gel
boxylic acid benzylamide (34).
flash chromatography on silica gel (hexanes/ethyl acetate, 1:1;
The product was purified by
(hexanes/ethyl acetate, 1:1; v/v) to afford the compound as white
crystals (73%) with mp 88.69 °C. 1H NMR (400 MHz, CDCl3): d 7.60

T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
643
v/v) to afford the compound as white crystals (97%) with mp 134–
136 °C. 1H NMR (400 MHz, CDCl3): d 7.44 (1H, dd, J = 0.7, 8.8),
7.40–7.35 (2H, m) 7.20–7.14 (3H, m), 6.85–6.80 (1H,ddd, J = 2.7,
5.1, 8.8), 6.80–6.69 (1H, ddd, J = 0.7, 2.7, 5.1), 5.15 (1H, dd, J = 2.3,
4.9), 4.34 (1H, dd, J = 1.0, 16.9),3.32 (1H, dd, J = 7.5, 16.0), 2.79
(1H, dd, J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3) d 165.24,
163.12, 162.51, 149.64, 139.30, 137.96, 128.84, 127.74, 112.53,
105.05, 56.59, 44.23, 43.84; IR (neat) mmax (C@O) 1774,
1709 cmÀ1. Anal. Calcd for C17H14F2N2O2S: C, 58.61; H, 4.05; N,
8.04. Found: C, 58.53; H, 4.06; N, 8.03.
5.1.3.9. 2-(3-Chloro-4-fluoro-phenylsulfanyl)-4-oxo-azetidine-
1-carboxylic acid benzylamide (38). The product is a yellow oil
purified by preparatory plate on silica gel (3:1 hexanes/ethyl ace-
tate) to give 37.5% yield. 1H NMR (400 MHz, CDCl3): d 7.45 (1H,
dd, J = 0.5, 2.6), 7.40–7.35 (2H, m), 7.30 (1H, dd, J = 2.5, 8.9),
7.15–7-10 (2H, m), 7.02 (1H, dd, J = 5.1, 8.7), 6.85 (1H, br s), 5.28
(1H, dd, J = 2.3, 5.0), 4.52 (2H, dd, J = 0.7, 17.0), 3.50 (1H, dd,
J = 7.5, 16.0), 2.93 (1H, dd, J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3)
d 165.06, 160.14, 157.63, 149.54, 137.65, 137.14, 135.20, 128.82,
127.73, 126.51, 121.85, 121.66, 117.42, 57.32, 44.19, 43.74,
30.98; IR (neat) mmax (C@O) 1774.60, 1704.76 cmÀ1. Anal. Calcd
for C17H14ClFN2O2S: C, 55.97; H, 3.87; Cl, 9.72; F, 5.21; N, 7.68;
O, 8.77; S, 8.79. Found: C, 56.08; H, 3.80; Cl, 9.51; F, 5.17; N,
7.61; O, 8.73; S, 8.57.
5.1.3.10.
2-(2,4-Dichloro-phenylsulfanyl)-4-oxo-azetidine-1-
carboxylic acid benzylamide (39). The product was purified by
flash chromatography on silica gel (hexanes/ethyl acetate, 1:1;
v/v) to afford the compound as white crystals (28%) with mp
95–97 °C. 1H NMR (400 MHz, CDCl3) dH 7.40 (1H, d, J = 2.2), 7.25
(1H, dd J = 2.2, 8.5), 7.13 (1H, dd, J = 0.6, 8.5), 7.0 (5H, m), 6.73
(1H, br s), 5.29 (1H, dd, J = 2.5, 4.8), 4.39 (2H, dd, J = 0.7, 15.0),
3.44 (1H, dd, J = 7.5, 16.0), 2.91 (1H, dd, J = 7.5, 16.0); 13C NMR
(100 MHz, CDCl3)
d 165.08, 149.50, 138.63, 137.65, 136.75,
135.77, 130.03, 128.91, 128.79, 127.86, 127.72, 56.62, 44.25,
43.74; IR (neat) mmax (C@O) 1774.53, 1705.07 cmÀ1. Anal. Calcd
for C17H14Cl2N2O2S: C, 53.55; H, 3.70; Cl, 18.60; N, 7.35; O, 8.39;
S, 8.41. Found: C, 53.78; H, 3.82; Cl, 18.57; N, 7.48; O, 8.31; S, 8.39.
5.1.3.6. N-Benzyl-2-((3,4-difluorophenyl)thio)-4-oxoazetidine-
1-carboxamide (35).
The product was purified by flash
5.1.3.11.
carboxamide (40).
N-Benzyl-2-((4-nitrophenyl)thio)-4-oxoazetidine-1-
The product was purified by flash chroma-
chromatography on silica gel (hexanes/ethyl acetate, 1:1; v/v) to
afford the compound as white crystals (97%) with mp 89.24 °C.
1H NMR (400 MHz, CDCl3): d 7.49–7.35 (2H, m), 7.23 (1H, ddd,
J = 0.6, 1.7, 5.1), 7.17–7.12 (4H, m) 6.90–7.10 (1H, ddd, J = 0.6, 5.1,
8.9), 6.75 (1H, br s), 5.17 (1H, dd, J = 2.3, 4.9), 4.43 (2H, dd J = 0.7,
17.0), 3.35 (1H, dd, J = 7.5, 16.0); 2.80 (1H, dd, J = 7.5, 15.7); 13C
NMR (100 MHz, CDCl3): d 165.28, 164.85, 162.36, 149.60, 137.76,
128.80, 127.74, 124.47, 116.55, 57.06, 43.70; IR (neat) mmax (C@O)
1775, 1700 cmÀ1. Anal. Calcd for C17H14F2N2O2S: C, 58.61; H,
4.05; N, 8.04. Found: C, 61.77; H, 4.49; N, 8.44.
tography on silica gel (hexanes/ethyl acetate, 1:1; v/v) to afford the
compound as white crystals (76%) with mp 123.04 °C.
1H NMR (400 MHz, CDCl3): d 8.40 (2H, dd, J = 2.0, 8.4), 7.65 (2H,
dd, J = 3.1, 8.5), 7.0 (5H, m), 6.82 (1H, br s), 5.42 (1H, dd, J = 2.3, 4.9),
4.42 (2H, dd, J = 0.7, 15.0), 3.59 (1H, dd J = 5.7, 16.0), 3.0 (1H, dd
J = 5.7, 16.0); 13C NMR (100 MHz, CDCl3)
d 164.94, 149.69,
147.22, 141.97, 137.61, 131.59, 129.00, 127.98, 127.84, 124.25,
56.80, 45.12, 43.98; IR (neat) mmax (C@O) 1775, 1700 cmÀ1. Anal.
Calcd for C17H15N3O4S: C, 57.13; H, 4.23; N, 11.76. Found: C,
57.43; H, 4.37; N, 12.01.
5.1.3.7. N-Benzyl-2-oxo-4-((perfluorophenyl)thio)azetidine-1-
carboxamide (36).
The product is a cloudy yellow oil purified
5.1.3.12. N-Benzyl-2-oxo-4-((2-(trifluoromethyl)phenyl)thio)-
azetidine-1-carboxamide (41). The product was purified by
flash chromatography on silica gel (hexanes/ethyl acetate, 1:1; v/
v) to afford the compound as viscous oil in 66% yield. 1H NMR
(400 MHz, CDCl3): d 7.84 (1H, d, J = 7.5), 7.67 (1H, d, J = 1.3, 7.6),
7.25 (2H, m); 7.0 (5H, m), 6.76 (1H, s), 5.28 (1H, dd, J = 2.3, 5.0),
4.37 (2H, dd J = 0.7, 15.0), 3.42 (1H, dd, J = 7.5, 16.0), 2.87 (1H,
dd, J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3) d 165.49, 149.77,
137.92, 137.69, 134.46 (1C, q, J = 0.3), 132.54, 130.49, 129.23,
128.95, 127.86, 127.81, 127.22 (1C, q, J = 0.05), 58.18, 44.62,
43.87; IR (neat) mmax (C@O) 1778, 1706 cmÀ1. Anal. Calcd for
by preparatory plate on silica gel (3:1 hexanes/ethyl acetate) to
give 37.5% yield. 1H NMR (400 MHz, CDCl3): d 7.39–7.46 (2H, m),
7.15–7.10 (3H. m) 6.69 (1H, br s), 5.46 (1H, dd J = 2.3, 4.9), 4.5
(1H, dd, J = 0.7, 17.0), 3.60 (1H, dd, J = 7.6, J = 16.0), 2.99 (1H, dd,
J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3)
d 164.11, 149.21,
128.78, 127.58, 77.36, 55.66, 43.76; IR (neat) mmax (C@O) 1775,
1700 cmÀ1. Anal. Calcd for C17H11F5N2O2S: C, 50.75; H, 2.76; N,
6.96. Found: C, 50.96; H, 2.93; N, 6.61.
5.1.3.8. 2-(2-Chloro-4-fluoro-phenylsulfanyl)-4-oxo-azetidine-
1-carboxylic acid benzylamide (37).
The product was
C18H15F3N2O2S: C, 56.84; H, 3.97; N, 7.36; Found: C, 56.92;
purified by flash chromatography on silica gel (hexanes/ethyl
acetate, 1:1; v/v) to afford the compound as white crystals (43%)
with mp116–119 °C. 1H NMR (400 MHz, CDCl3): d 7.40–7.35 (2H,
dd, J = 1.4, 7.6), 7.26 (1H, d, J = 8.8),7.15–7.10 (3H, m) 6.97
(1H, dd, J = 2.8, 5.1), 6.87 (1H, dd, J = 5.1, 8.8), 6.82 (1H, br s),
5.36 (1H, dd, J = 2.3, 5.0), 3.48 (1H, dd, J = 0.7, 17.0), 3.51 (1H, dd,
J = 7.5, 16.0), 2.98 (1H, dd, J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3)
d 165.14, 149.51, 138.62, 137.69, 128.78, 127.71, 117.92, 114.98,
H, 4.03; N, 7.45.
5.1.3.13. N-Benzyl-2-oxo-4-((3-(trifluoromethyl)phenyl)thio)-
azetidine-1-carboxamide (42).
The product was purified by
flash chromatography on silica gel (hexanes/ethyl acetate, 1:1;
v/v) to afford the compound as viscous oil in 68% yield. 1H NMR
(400 MHz, CDCl3): d 7.77 (1H, m), 7.74 (1H, ddd, J = 1.6, 1.8, 7.9),
7.54 (2H, ddd, J = 1.0, 7.7, 7.9), 7.18–7.29 (2H, m), 6.72 (1H, br s),
5.26 (1H, dd, J = 2.3, 4.9), 4.41 (1H, dd, J = 0.7, 15.0), 3.41 (1H, dd,
J = 7.5, 16.0), 2.82 (1H, dd, J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3)
d 165.20, 149.71, 137.79, 137.74, 132.13, 44.57, 131.74 (1C, q,
J = 0.32), 131.14, 129.88, 128.95, 127.88, 127.85, 125.90, 57.21,
56.91, 44.01, 44.73; IR (neat) mmax (C@O) 1772.41, 1700.35 cmÀ1
.
Anal. Calcd for C17H14ClFN2O2S: C, 55.97; H, 3.87; Cl, 9.72; F,
5.21; N, 7.68; O, 8.77; S, 8.79. Found: C, 56.17; H, 4.07; Cl, 9.81;
F, 5.19; N, 7.83; O, 8.68; S, 8.67.

644
T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
43.91; IR (neat) mmax (C@O) 1777, 1706 cmÀ1. Anal. Calcd for C18-
H15F3N2O2S: C, 56.84; H, 3.97; N, 7.36; Found: C, 57.01; H, 4.02;
N, 7.46.
Calcd for C18H18N2O3S: C, 63.14; H, 5.30; N, 8.18. Found: C,
63.34; H, 5.13; N, 8.18.
5.1.3.19. N-Benzyl-2-((2,5-dimethoxyphenyl)thio)-4-oxoazeti-
dine-1-carboxamide (48). The compound was purified by recrys-
tallization in hexanes/ethyl acetate to give a white powder (88%)
with mp 87.5–88.5 °C. 1H NMR (400 MHz, CDCl3): d 7.48 (5H, m),
6.87 (2H, m), 6.74 (1H, d, J = 2.42), 5.21 (1H, dd, J = 2.6, 5.0), 4.27
(2H, m), 3.75 (3H, s), 3.75 (3H, s), 3.36 (1H, dd, J = 7.5, 16.0), 2.86
(1H, dd, J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3) d 128.75,
128.63, 127.74, 127.64, 127.49, 121.90, 116.30, 112.16, 56.35,
55.77, 44.04, 43.67; IR (neat) mmax (C@O) 1775, 1705 cmÀ1. Anal.
Calcd for C19H20N2O4S: C, 61.27; H, 5.41; N, 7.52. Found: C,
61.04; H, 5.39; N, 7.47.
5.1.3.14. N-Benzyl-2-oxo-4-((4-(trifluoromethyl)phenyl)thio)-
azetidine-1-carboxamide (43).
The product was purified by
flash chromatography on silica gel (hexanes/ethyl acetate, 1:1;
v/v) to afford the compound as white crystals (64%) with
mp72–73 °C. 1H NMR (400 MHz, CDCl3): d 7.67 (1H, dd, J = 2.0,
8.2), 7.60 (1H, dd, J = 1.9, 8.3), 7.39 (5H, m), 6.87 (1H,br s), 5.41
(1H, dd, J = 2.3, 4.9), 4.52 (2H, dd, J = 0.7, 15.0), 3.57 (1H, dd,
J = 7.5, 16.0), 2.99 (1H, dd, J = 7.9, 16.0); 13C NMR (100 MHz, CDCl3)
d 165.26, 149.75, 137.81, 136.34, 133.38, 30.62 (1C, q, J = 0.33),
127.87, 126.20, 57.01, 44.86, 127.93, 128.98, 143.91; IR (neat) mmax
(C@O) 1778, 1708 cmÀ1. Anal. Calcd for C18H15F3N2O2S: C, 56.84; H,
3.97; N, 7.36; Found: C, 56.98; H, 4.01; N, 7.39.
5.1.3.20. N-Benzyl-2-((3,4-dimethoxyphenyl)thio)-4-oxoazeti-
dine-1-carboxamide (49). The compound was purified by prepara-
tory plate on silica gel (3:1 hexanes/ethyl acetate) to give a clear oil
in 63% yield. 1H NMR (400 MHz, CDCl3) dH 7.48–6.84 (4H, m), 5.21
(1H, dd, J = 2.65, 2.91), 3.82 (3H, s), 3.36 (1H, dd, J = 5.61, 10.69),
2.86 (1H, dd, J = 2.67, 13.65); 13C NMR (100 MHz, CDCl3) d 165.59,
150.38, 149.68, 149.03, 137.93, 129.29, 128.73, 127.75, 127.64,
119.13, 118.98, 118.77, 111.37, 56.74, 55.88, 43.58, 43.37; IR (neat)
5.1.3.15. N-Benzyl-2-((3,5-bis(trifluoromethyl)phenyl)thio)-4-
oxoazetidine-1-carboxamide (44).
The product was purified
by flash chromatography on silica gel (hexanes/ethyl acetate, 1:1;
v/v) to afford the compound as white crystals (65%) with mp 61–
62 °C. 1H NMR (400 MHz, CDCl3): d 8.05 (1H, m), 7.77 (1H, m),
7.19–7.30 (5H, m), 6.72 (1H, br s), 5.33 (1H, dd, J = 2.3, 4.9),
4.36–4.46 (2H, m), 3.53 (1H, dd, J = 7.5, 16.0), 2.91 (1H, dd,
m
max (C@O) 1775, 1705 cmÀ1. Anal. Calcd for C19H20N2O4S: C, 61.27;
J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3)
d
164.80, 149.62,
H, 5.41; N, 7.52. Found: C, 61.54; H, 5.56; N, 7.43.
137.61, 135.40, 133.26, 132.52 (2C, q, J = 0.33), 128.97, 127.94,
127.88, 124.40, 122.45, 121.68, 57.49, 44.83, 43.99; IR (neat) mmax
(C@O) 1780, 1707 cmÀ1. Anal. Calcd for C19H14F6N2O2S: C, 50.89;
H, 3.15; N, 6.25; Found: C, 50.93; H, 3.42; N, 6.41.
5.1.3.21. N-Benzyl-2-oxo-4-(thiophen-3-ylthio)azetidine-1-car-
boxamide (50).
The compound was purified by preparatory
plate on silica gel (3:1 hexanes/ethyl acetate) to give a clear oil
72% yield. 1H NMR (400 MHz, CDCl3): d 7.35 (1H, d, J = 5.2), 7.12
(1H, dd, J = 3.7, 5.2), 6.95 (5H, m), 6.75 (1H, dd, J = 1.3, 3.6), 6.68
(1H, br s), 5.05 (1H, dd, J = 2.3, 4.9), 4.42 (2H, m), 3.25 (1H, dd,
J = 7.5, 16.0), 2.85 (1H, dd, J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3)
d 165.23, 149.48, 138.16, 137.78, 132.52, 128.15, 128.75, 127.77,
127.69, 125.34, 56.49, 43.71, 43.11, 30.98, 29.73; IR (neat) mmax
(C@O) 1770, 1700 cmÀ1. Anal. Calcd for C15H14N2O2S2: C, 56.58;
H, 4.43; N, 8.80. Found: C, 56.72; H, 4.65; N, 8.92.
5.1.3.16. N-Benzyl-2-((2-methoxyphenyl)thio)-4-oxoazetidine-
1-carboxamide (45).
The product was purified by recrystalli-
zation in hexane/ethyl acetate to give a white powder (90%) with
mp 105–107 °C. 1H NMR (400 MHz, CDCl3): d 7.43 (1H, dd,
J = 1.7, 15.0), 7.20 (1H, ddd, J = 1.7, 7.2, 8.0), 7.0 (5H, m), 6.87
(1H, ddd, J = 1.3, 7.2, 9.1), 6.68 (1H, dd, J = 1.3, 8.0), 5.38 (1H, dd,
J = 2.3, 4.9), 4.26 (2H, m), 3.86 (3H, s), 3.40 (1H, dd, J = 7.5, 16.0),
2.94 (1H, dd, J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3) d 165.70,
159.93, 149.61, 137.88, 137.28, 131.25, 128.74, 127.75, 127.64,
121.25, 117.37, 111.22, 55.81, 43.94, 43.65; IR (neat) mmax (C@O)
1627, 1576 cmÀ1. Anal. Calcd for C18H18N2O3S: C, 63.14; H, 5.30;
N, 8.18. Found: C, 63.40; H, 5.28; N, 8.32.
5.1.3.22. N-Benzyl-2-((4-(methylthio)phenyl)thio)-4-oxoazeti-
dine-1-carboxamide (51).
The compound was purified by
preparatory plate on silica gel (3:1 hexanes/ethyl acetate) to give
a clear oil in 66% yield. 1H NMR (400 MHz, CDCl3) dH 7.47 (2H, d,
J = 8.1), 7.15 (2H, d, J = 8.1), 7.0 (5H, m), 6.84 (1H, br s), 5.26 (1H,
dd, J = 2.3, 5.0), 4.53 (2H, dd, J = 0.7, 15.0), 3.42 (1H, dd, J = 7.5,
16.0), 2.89 (1H, dd, J = 7.5, 16.0), 2.49 (3H, s); 13C NMR (100 MHz,
CDCl3) d 165.43, 149.65, 141.11, 137.99, 135.93, 128.77, 127.67,
124.67, 56.76, 43.79, 43.64, 15.20; IR (neat) mmax (C@O) 1700,
1780 cmÀ1. Anal. Calcd for C18H18N2O2S: C, 60.31; H, 5.06; N,
7.81. Found: C, 60.50; H, 5.05; N, 7.89.
5.1.3.17. N-Benzyl-2-((3-methoxyphenyl)thio)-4-oxoazetidine-
1-carboxamide (46).
The product purified via a flash column
chromatography on silica gel with a ratio gradient of 3:1 hexanes
and dichloromethane to produce a colorless oil with a 57% yield.
1H NMR (400 MHz, CDCl3): d 7.25–7.33 (3H, m), 7.7 (1H, ddd,
J = 1.7, 2.1, 8.1), 7.0 (5H, m), 5.19 (1H, dd, J = 2.3, 4.9), 3.68 (3H,
s), 3.22 (1H, dd, J = 7.0, 16.0), 2.78 (1H, dd, J = 7.5, 16.0); 13C NMR
5.1.3.23. N-Benzyl-2-(naphthalen-1-ylthio)-4-oxoazetidine-1-
(100 MHz, CDCl3)
d 165.53, 159.87, 149.63, 137.79, 130.47,
carboxamide (52).
The compound was purified by prepara-
130.04, 128.78, 127.74, 127.13, 120.01, 115.34, 56.60, 55.35,
44.01, 43.69; IR (neat) mmax (C@O) 1775, 1700 cmÀ1. Anal. Calcd
for C18H18N2O3S: C, 63.14; H, 5.30; N, 8.18. Found: C, 63.57; H,
5.51; N, 8.05.
tory plate on silica gel (3:1 hexanes/ethyl acetate) to give an opaque
oil in 73% yield. 1H NMR (400 MHz, CDCl3): d 7.80–7.50 (7H, m), 7.0
(5H, m), 6.87 (1H, br s), 5.40 (1H, dd, J = 2.3, 4.9), 4.56 (2H, dd,
J = 1.0, 15.0), 3.42 (1H, dd, J = 7.5, 16.0), 2.95 (1H, dd, J = 7.5, 15.0);
13C NMR (100 MHz, CDCl3): d 165.42, 149.70, 135.11, 137.92,
133.54, 133.20, 131.55, 128.96, 128.82, 127.86, 127.81, 127.78,
127.71, 127.16, 126.80, 126.63, 56.72, 44.09, 43.71, 29.76; IR (neat)
5.1.3.18. N-Benzyl-2-((4-methoxyphenyl)thio)-4-oxoazetidine-
1-carboxamide (47).
The compound was purified by recrys-
m
max (C@O) 1780, 1700 cmÀ1. Anal. Calcd for C21H18N2O2S: C, 69.59;
H, 5.01; N, 7.73; Found: C, 69.79; H, 5.30; N, 7.55.
tallization in hexanes/ethyl acetate to give a white powder (58%)
with mp 105–110 °C. 1H NMR (400 MHz, CDCl3): d 7.48 (2H, d,
J = 9.0), 7.2 (2H, d, J = 8.9), 6.98 (5H, m), 5.21 (1H, dd, J = 2.3, 4.9),
4.27 (2H, m), 3.82 (3H, s), 3.36 (1H, dd, J = 7.5, 16.0), 2.86 (1H,
dd, J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3) d 194.66, 165.53,
160.88, 137.95, 128.76, 127.78, 127.67, 118.74, 114.83, 56.72,
55.35, 43.64, 43.39; IR (neat) mmax (C@O) 1775, 1700 cmÀ1. Anal.
5.1.3.24.
boxamide (53).
N-Benzyl-2-(4-nitrophenoxy)-4-oxoazetidine-1-car-
The compound was purified by recrystalliza-
tion in hexanes/ethyl acetate to give yellow crystals in 72% yield
with mp 123–124 °C. 1H NMR (400 MHz, CDCl3): d 8.28 (2H, d,

T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
645
J = 9.2); 7.58 (2H, d, J = 9.3), 7.0–7.1 (5H, m), 6.86 (1H, br s), 6.18 (1H,
dd, J = 1.5, 3.6), 4.51 (2H, d, J = 0.7, 15.0), 3.58 (1H, dd, J = 1.5, 16.0),
3.25 (1H, d, J = 1.5, 16.2); 13C NMR (100 MHz, CDCl3) d 164.73,
161.08, 149.38, 143.04, 137.31, 128.84, 127.84, 127.69, 125.89,
5.1.3.30. 2-Oxo-4-phenylsulfanyl-azetidine-1-carboxylic acid
benzhydryl-amide (59). To a solution of 2 (0.107 g, 0.596
mmol) and diphenylmethyl isocyanate (0.116 ml, 0.615 mmol) in
methylene chloride (5 ml) was added triethylamine (85.5 L,
l
116.65, 78.12, 45.54, 43.84; IR (neat) mmax (C@O) 1715, 1775 cmÀ1
.
0.614 mmol). The solution was microwave irradiated under pres-
sure at 70 °C for 20 min. The solution was washed with 5% HCl
(10 ml aliquots, 3 times) and the product was purified by column
chromatography with a linear mobile phase gradient (10:1 methy-
lene chloride/ethyl acetate to 1:10 methylene chloride/ethyl ace-
tate). This procedure yielded product 7 (0.19 g, 86.3%) as a light
brown gel. 1H NMR (400 MHz, CDCl3): d 7.54 (2H, dt, J = 1.3, 7.2),
7.35 (5H, m), 7.27 (4H, dt, J = 2.4, 7.4), 7.21 (4H, dd, J = 1.3, 7.4),
7.07 (1H, m), 5.99 (1H, br s), 5.29 (1H, dd, J = 2.3, 4.9), 3.44 (1H, dd,
J = 7.5, 15.9), 2.91 (1H, dd, J = 7.5, 16.0).
Anal. Calcd for C17H15N3O5: C, 59.82; H, 4.43; N, 12.31. Found: C,
59.67; H, 4.53; N, 12.20.
5.1.3.25. N-Benzyl-2-oxo-4-(4-(trifluoromethyl)phenoxy)azeti-
dine-1-carboxamide (54).
The compound was purified by
recrystallization in hexanes/ethyl acetate to give a white powder
in 70% yield with mp 103–109 °C. 1H NMR (400 MHz, CDCl3): d
7.54 (2H, d, J = 8.5), 7.17–7.31 (7H, m), 6.79 (1H, br s), 6.04 (1H,
dd, J = 1.5, 3.6), 4.43 (2H, d, J = 1.0, 15.0), 3.44 (1H, dd, J = 1.5,
16.0), 3.11 (1H, dd, J = 1.6, 16.0); 13C NMR (100 MHz, CDCl3) d
165.05, 158.68, 149.47, 137.42, 128.82, 127.78, 127.69, 127.19,
127.08, 125.34, 122.78, 116.76, 78.27, 45.52, 43.80; IR (neat) mmax
(C@O) 1690, 1780 cmÀ1. Anal. Calcd for C18H15F3N2O3: C, 59.34;
H, 4.15; N, 7.69. Found: C, 59.31; H, 4.37; N, 7.63.
5.1.3.31. 2-Oxo-4-phenylsulfanyl-azetidine-1-carboxylic acid
(9H-fluoren-9-yl)-amide (60).
0.664 mmol) and 9H-fluoren-9-yl
0.798 mmol) in methylene chloride (5 ml) was added triethyl-
amine (95.3 L, 0.683 mmol). The solution was microwave irradi-
To a solution of 2 (0.119 g,
isocyanate (0.141 g,
l
5.1.3.26. N-Benzyl-2-oxo-4-(phenylselanyl)azetidine-1-carbox-
ated under pressure at 70 °C for 15 min. The solution was
washed with 5% HCl (10 ml aliquots, 3 times) and the product
was purified by column chromatography with a linear mobile
phase gradient (10:1 methylene chloride/ethyl acetate to 1:10
methylene chloride/ethyl acetate). This procedure yielded the
product as a yellow solid (0.22 g, 89.7%). 1H NMR (400 MHz,
CDCl3): d 7.80 (2H, dd, J = 1.4, 7.7), 7.36–7.31 (4H, m), 7.30–7.24
(5H, m), 7.18 (1H, m), 7.07 (1H, m), 6.92 (2H, ddd, J = 1.4, 7.3,
7.7), 5.18 (1H, dd, J = 2.3, 4.9), 3.19 (1H, dd, J = 7.5, 16), 2.67 (1H,
dd, J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3) d 148.16, 141.66,
141.36, 138.31, 133.07, 127.14, 127.05, 126.69, 125.63, 122.86,
amide (55).
The compound was purified by recrystallization
in hexanes/ethyl acetate to give a white powder in 30% yield. 1H
NMR (400 MHz, CDCl3): d 7.35–7.89 (5H, m), 7.0 (5 h, m), 6.33
(1H, br s), 5.14 (1H, dd, J = 2.3, 4.9), 4.27 (2H, m), 3.45 (1H, ddd,
J = 1.7, 7.8, 14.8), 2.97 (1H, ddd, J = 1.6, 7.8, 15.6).
13C NMR (100 MHz, CDCl3) d 165.49, 149.60, 137.84, 136.83,
129.35, 128.78, 127.72, 127.32, 126.32, 125.05, 48.15, 44.66,
43.69, 14.24; IR (neat) mmax (C@O) 1771.21, 1701.02 cmÀ1. Anal.
Calcd for C17H16N2O2Se: C, 56.83; H, 4.49; N, 7.80; O, 8.91; Se,
21.98. Found: C, 56.97; H, 4.69; N, 7.73; O, 8.74; Se, 22.07.
117.89, 54.56, 52.55, 41.81. Anal. Calcd for
C23H18N2O2S:
5.1.3.27. N-Benzyl-2-oxoazetidine-1-carboxamide (56).
The
C, 71.48; H, 4.69; N, 7.25; O, 8.28; S, 8.30. Found: C, 71.64; H,
4.68; N, 7.43; O, 8.47; S, 8.51.
synthesis was performed as developed in our laboratory. To a solu-
tion of the appropriate azetidin-2-one 5 (0.1 g, 0.46 mmol in
dichloromethane; 4 ml), was added 1.2 mol equiv of triethylamine
and 1.1 mol equiv of the corresponding isocyanate (2-fluorobenzyl
isocyanate, 2-fluorophenyl isocyanate, or 2-methoxybenzyl isocya-
nate). The reaction was irradiated under pressure (CEM micro-
wave, 10–60 min, 300 W, 35 °C) and monitored by TLC and NMR.
After 10 min of irradiation, the product was confirmed by NMR.
Upon evaporation the product proved unstable within 24 h. 1H
NMR (400 MHz, CDCl3): d 7.13–7.28 (1H, m), 4.39 (1H, d, J = 6.0),
3.55 (1H, t, J = 4.7), 2.95 (1H, t, J = 4.7); 13C NMR (100 MHz, CDCl3)
d 167.05, 150.62, 137.94, 128.50, 128.21, 43.68, 37.23, 36.07,
5.1.3.32. 2-Oxo-4-phenylsulfanyl-azetidine-1-carboxylic acid
naphthalen-1-ylamide (61).
To a solution of 2 (0.139 g,
0.775 mmol) and 1-naphthyl isocyanate (0.115 ml, 0.679 mmol)
in methylene chloride (5 ml) was added triethylamine (0.111 ml,
1.09 mmol). The solution was irradiated using CEM microwave at
70° C for 20 min. The solution was then washed with 5% HCl
(10 ml aliquots, 3 times). The resulting organic layer was evapo-
rated then purified by column chromatography with a linear
mobile phase gradient (10:1 methylene chloride/ethyl acetate to
1:10 methylene chloride/ethyl acetate). This procedure yielded
the product as a clear, colorless gel (0.21 g, 82.5%). 1H NMR
(400 MHz, CDCl3) dH 8.02 (2H, dd, J = 1.1, 8.0), 7.92 (1H, d, J = 1.5,
8.1), 7.71 (2H, dd, J = 1,2, 8.1), 7.54 (1H, dd, J = 1.5, 8.0), 7.51 (2H,
dd, J = 1.5, 8.0), 7.34 (2H, dt, J = 1.5, 7.6), 7.25 (2H, m), 7.23 (2H,
d, J = 8.0), 5.25 (1H, dd, J = 2.3, 4.9), 3.46 (1H, dd, J = 7.5, 16.0),
2.91 (1H, dd, J = 7.5. 16.0); 13C NMR (100 MHz, CDCl3) d 166.30,
147.45, 135.33, 134.01, 131.69, 129.44, 129.34, 128.83, 126.66,
126.18, 125.86, 125.73, 125.20, 120.14, 118.59, 57.24, 44.15. Anal.
Calcd for C20H16N2O2S: C, 68.95; H, 4.63; N, 8.04; O, 9.18; S, 9.20.
Found: C, 69.07; H, 4.78; N, 7.94; O, 9.11; S, 9.12.
30.07; IR (neat) mmax (C@O) 1760, 1700 cmÀ1
.
5.1.3.28. 2-(Allylthio)-N-benzyl-4-oxoazetidine-1-carboxamide
(57). The compound yield obtained was 33%; however, it was
unstable and appears to undergo a [2+2] cycloreversion. 1H NMR
(400 MHz, CDCl3): d 7.27–7.36 (9H, m), 6.90 (1H, br s), 5.88 (1H,
m), 5.33 (2H, d, J = 16.0), 5.19 (1H, d, J = 9.9), 4.50 (2H, s), 3.92 (1H,
dd, J = 4.0, 9.0), 3.42 (2H, ddd, J = 10.1, 5.3, 2.0), 2.92 (1H, d, J = 16.3).
5.1.3.29. 1-Hex-5-ynoyl-4-phenylsulfanyl-azetidin-2-one (58). The
compound was purified by extraction with hexanes from the reac-
tion mixture to give yellow oil in 10% yield. However, it was prone
to hydrolysis on silicagel.
5.1.3.33.
dine-1-carboxamide (62).
N-Benzhydryl-2-((4-fluorophenyl)thio)-4-oxoazeti-
The compound was purified by
1H NMR (400 MHz, CDCl3): d 7.36–7.56 (6H, m), 5.25 (2H, dd,
J = 2.9, 3. 2), 3.43 (2H, dd, J = 6.1, 10.5), 2.93 (2H, dd, J = 13.5, 3.2),
2.85 (2H, dt, J = 4.4, 2.8), 2.60 (1H, t, J = 7.2), 2.54 (2H, t, J = 7.4),
2.19–2.34 (1H, m), 2.25 (1H, dt, J = 2.5, 4.2), 2.01 (2H, t, J = 2.6),
1.92 (2H, dt, J = 1.6, 5.5).
preparatory plate on silica gel (3:1 hexanes/ethyl acetate) to give a
waxy solid in 28% yield. 1H NMR (400 MHz, CDCl3): d 7.55 (2H, dt,
J = 1.3, 7.1), 7.36 (2H, dd, J = 2.0, 8.9), 7.25 (4H, m), 7.23 (4H, d,
J = 8.0), 6.87 (1H, m), 6.41 (1H, br s), 5.09 (1H, dd, J = 2.3, 4.9), 3.50
(1H, dd, J = 7.5, 16.0), 3.13 (1H, dd, J = 7.5, 16.0); 13C NMR
(100 MHz, CDCl3) d 165.48, 164.87, 162.38, 148.82, 141.31, 140.93,
137.91, 128.80, 127.76, 127.28, 124.20, 116.48, 60.44, 57.15, 43.99,
13C NMR (100 MHz, CDCl3) d 169.78, 163.40, 135.19, 129.40,
127.30, 83.18, 69.34, 55.60, 43.76, 35.29, 31.00, 23.30, 22.49,
22.20, 17.77; IR (neat) mmax (C@O) 1700, 1795 cmÀ1
.
29.75; IR (neat) m
max (C@O) 1770.42, 1708.96 cmÀ1. Anal. Calcd for

646
T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
C23H19FN2O2S: C, 67.96; H, 4.71; F, 4.67; N, 6.89; O, 7.87; S, 7.89.
Found: C, 68.16; H, 4.81; F, 4.65; N, 6.95; O, 7.73; S, 7.79.
acetate) of the filtrate to give a waxy solid in 34.3% yield. 1H
NMR (400 MHz, CDCl3): d 7.44 (2H, ddd, J = 1.80, 7.6, 8.4), 7.03
(3H, m), 6.83 (1H, ddd, J = 2.7, 5.1, 8.8), 6.76 (1H, dd, J = 2.7, 5.1),
5.22 (1H, dd, J = 2.3, 4.9), 4.45 (1H, dd, J = 0.7, 15.0), 3.47
(1H, dd, J = 7.5, 16.0), 2.83 (1H, dd, J = 7.5, 15.0); 13C NMR
5.1.3.34. N-(2,2-Diphenylethyl)-2-((4-fluorophenyl)thio)-4-oxo-
azetidine-1-carboxamide (63). The compound was purified by
preparatory plate on silica gel (3:1 hexanes/ethyl acetate) to give a
solid in 32% yield. mp 155–156 °C. 1H NMR (400 MHz, CDCl3): d 7.34
(6H, m), 7.27–7.23 (2H, m), 7.15 (4H, dt, J = 1.3, 7.3), 6.95 (2H, dd,
J = 5.5, 8.9) 6.41 (1H, br s), 5.09 (1H, dd, J = 2.3, 4.9), 3.80 (2H, dd,
J = 7.3), 3.76 (1H, dt, J = 0.7, 7.3), 3.45 (1H, dd, J = 7.5, 16.0), 3.15 (1H,
dd, J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3) d 165.00, 141.54, 141.41,
137.85, 128.80, 128.08, 126.99, 124.11, 116.59, 116.37, 56.61, 50.87,
(100 MHz, CDCl3)
d 166.05, 162.37, 159.41, 157.24, 151.09,
131.35, 130.19, 129.60, 128.90, 124.96, 122.07, 115.43,
111.27, 105.02, 63.30, 43.01, 38.95. IR (neat) mmax (C@O) 1774,
1709 cmÀ1. Anal. Calcd for C17H13F3N2O2S: C, 55.73; H, 3.58; F,
15.56; N, 7.65; O, 8.73; S, 8.75. Found: C, 55.93; H, 3.72; F, 15.67;
N, 7.78; O, 8.79; S, 8.59.
44.07, 43.45; IR (neat) m
max (C@O) 1772.23, 1705.13 cmÀ1. Anal. Calcd
for C24H21FN2O2S: C, 68.55; H, 5.03; F, 4.52; N, 6.66; O, 7.61; S, 7.62.
Found: C, 68.72; H, 5.24; F, 4.63; N, 6.67; O, 7.52; S, 7.83.
5.1.3.38. 2-((2,4-Difluorophenyl)thio)-N-(2-methoxybenzyl)-4-
oxoazetidine-1-carboxamide (67).
The compound was syn-
thesized under the conditions as stated above for compound 61
and purified by preparatory place on silica gel (1:1 hexanes/ethyl
acetate) of the filtrate to give a waxy solid in 62.6% yield. 1H
NMR (400 MHz, CDCl3): d 7.47, (1H, d, J = 8.8), 7.39 (1H, ddd,
J = 1.7, 7.7, 8.2), 7.17–7.04 (3H, m), 6.82 (1H, ddd, J = 2.7, 5.1,
8.8), 6.74 (1H, dt, J = 2.7, 5.1) 5.15 (1H, dd, J = 2.3, 4.9), 4.38
(1H, dd, J = 0.7, 15.0), 3.78 (1H, s), 3.35 (1H, dd J = 7.5, 16.0),
2.79 (1H, dd, J = 7.5, 16.0); 13C NMR (100 MHz, CDCl3)
d
165.35, 164.87, 162.89, 162.24, 157.59, 149.35, 139.19, 129.60,
129.10, 125.86, 120.57, 112.43, 112.23, 110.32, 105.06, 104.80,
104.53, 56.30, 55.28, 39.75, 30.07; IR (neat) mmax (C@O) 1775,
1700 cmÀ1. Anal. Calcd for C18H16F2N2O3S: C, 57.14; H, 4.26; F,
10.04; N, 7.40; O, 12.68; S, 8.47. Found: C, 57.34; H, 4.32; F,
10.21; N, 7.51; O, 12.49; S, 8.37.
5.1.3.39.
2-((2,4-Difluorophenyl)thio)-N-(2-fluorophenyl)-4-
The compound was syn-
oxoazetidine-1-carboxamide (68).
thesized under the conditions as stated above for compound 61.
The product was confirmed by NMR, but the product proved
unstable upon evaporation of the solvent.
5.1.3.35. 2-((2,4-Difluorophenyl)thio)-N-ethyl-4-oxoazetidine-
1-carboxamide (64). The compound was purified by flash chro-
matography on silica gel with 2:1 hexanes/ethyl acetate to give a
waxy solid in 95% yield. 1H NMR (400 MHz, CDCl3): d 7.63 (1H,
dd, J = 0.5, 8.80), 7.25 (1H, ddd, J = 2.7, 5.1, 8.8), 6.92 (1H, dt,
J = 0.5, 2.7, 5.2), 6.39 (1H, br s), 5.26 (1H, dd J = 2.3, 4.9), 3.46
(1H, dd, J = 7.5, 16.0), 3.33 (1H, dd, J = 7.5, 16.0), 2.90 (1H, q,
J = 7.35, 14.0), 1.20 (1H, t, J = 7.3);13C NMR (100 MHz, CDCl3) d
165.10, 149.4, 138.91, 112.45, 104.89, 77.05, 43.99, 34.75, 15.08;
IR (neat) mmax (C@O) 1775 cmÀ1. Anal. Calcd for C12H12F2N2O2S:
C, 50.34; H, 4.22; F, 13.27; N, 9.78; O, 11.18; S, 11.20. Found:
C, 50.61; H, 4.34; F, 13.18; N, 9.87; O, 11.23; S, 11.13.
1H NMR (400 MHz, CDCl3): d 7.82 (1H, ddd, J = 1.4, 5.1, 8.4), 7.79
(1H, dd, J = 1.4, 8.0), 6.99 (1H, ddd, J = 1.4, 7.8, 8.4), 6.82 (1H, ddd,
J = 2.7, 5.1, 8.8), 6.75 (1H, dt, J = 2.7, 5.1), 5.28 (1H, dd, J = 2.3,
4.9), 3.47 (1H, dd, J = 7.5, 16.0), 2.90 (1H, dd, J = 7.5, 16.0).
5.1.3.40. 2-((2,4-Difluorophenyl)thio)-N-(2-nitrobenzyl)-4-oxo-
azetidine-1-carboxamide (69). The synthesis was performed as
described by Mulchande et al.23 with the following modifications:
0.04 mmol, 10 mg of lactam 6 were dissolved in CDCl3 (1 ml)
with 1.5 mol equiv of 2-nitrophenyl isocyanate. Triethylamine
(0.001214 g; 0.3 mol equiv) was then added dropwise. The reaction
was stirred, and monitored by NMR. It was confirmed the product
was present after 30 min. The product became unstable with
removal of the solvent. 1H NMR (400 MHz, CDCl3): d 8.98 (1H,
dd, J = 1.4, 7.9), 8.06 (1H, dt, J = 1.4, 7.4), 7.65 (1H, d, J = 8.8), 7.54
(1H, dd, J = 1.4, 7.4), 7.34 (1H, ddd, J = 1.4, 7.5, 8.0), 6.84 (1H, ddd,
J = 2.7, 5.1, 8.8), 6.73 (1H, dt, J = 2.7, 5.1), 6.02 (1H, dd, J = 2.3,
4.9), 3.39 (1H, dd, J = 7.5, 16.0), 2.82 (1H, dd, J = 7.5, 16.0).
5.1.3.36.
N-(1-(Naphthalen-1-yl)ethyl)-2-oxo-4-(phenylthio)-
azetidine-1-carboxamide (65). The compound was purified by
flash chromatography on silica gel with 2:1 hexanes/ethyl acetate to
give a waxy solid in 55% yield. 1H NMR (400 MHz, CDCl3): d 8.05 (1H,
dd, J = 1.4, 8.0), 7.93 (1H, dd, J = 1.5, 8.4), 7.71 (1H, dd, J = 1.5, 7.9),
7.62 (1H, dd, J = 1.4, 7.9), 7.60 (1H, dd, J = 0.8, 7.7), 7.47 (1H, d,
J = 8.8), 7.43 (1H, tq, J = 7.3, 7.8), 7.31 (1H, dd, J = 1.4, 7.1), 6.82 (1H,
ddd, J = 2.7, 5.2, 8.8), 6.75 (1H, tq, J = 2.7, 5.2), 6.38 (1H, br s), 5.86
(1H, dd, J = 2.3, 4.9), 5.05 (1H, q, J = 6.9), 3.23 (1H, dd, J = 7.5, 16.0),
2.72 (1H, dd, J = 7.5, 16.0),1.57 (3H, d, J = 6.9); 13C NMR (100 MHz,
CDCl3) d 165.36, 148.71, 138.17, 137.52, 133.95, 130.67, 128.93,
128.37, 126.52, 125.82, 125.38, 124.92, 123.05, 122.46, 116.52, 57.30,
5.1.3.41.
N-Benzhydryl-2-((2-chloro-4-fluorophenyl)thio)-4-
The compound was syn-
oxoazetidine-1-carboxamide (70).
thesized under the conditions as stated above for compound 61
and purified by preparatory place on silica gel (1:1 hexanes/ethyl
acetate) of the filtrate to give a waxy solid in 30% yield. 1H NMR
(400 MHz, CDCl3): d 7.56 (2H, dt, J = 1.3, 7.2), 7.21 (2H, dt, J = 1.4,
7.1), 7.16 (2H, ddd, J = 1.2, 1.4, 7.4), 6.97 (1H, dd, J = 2.6, 5.1),
6.87 (1H, ddd, J = 2.5, 5.1, 8.8), 6.12 (1H, s), 5.25 (1H, dd, J = 2.3,
4.9), 3.42 (1H, dd, J = 7.5, 16.0), 2.90 (1H, dd, J = 7.5, 16.0); 13C
NMR (100 MHz, CDCl3) d 165.36, 164.36, 161.83, 148.72, 140.97,
140.10, 138.79, 128.80, 127.73, 127.32, 125.03, 117.92, 114.98,
45.49, 44.08, 22.05; IR (neat) m
max (C@O) 1772.15, 1700.24 cmÀ1. Anal.
Calcd for C22H19FN2O2S: C, 66.99; H, 4.86; F, 4.82; N, 7.10; O, 8.11; S,
8.13. Found: C, 67.19; H, 4.95; F, 4.79; N, 7.22; O, 8.12; S, 8.10.
5.1.3.37. 2-((2,4-Difluorophenyl)thio)-N-(2-fluorobenzyl)-4-ox-
oazetidine-1-carboxamide (66). The compound was synthe-
sized under the conditions as stated above for compound 61 and
purified by preparatory place on silica gel (2:1 hexanes/ethyl
57.29, 56.97, 44.08. IR (neat) mmax (C@O) 1772.09, 1716.36 cmÀ1
.

T. N. Beck et al. / Bioorg. Med. Chem. 23 (2015) 632–647
647
Anal. Calcd for C23H18ClFN2O2S: C, 62.65; H, 4.12; Cl, 8.04; F, 4.31;
References and notes
N, 6.35; O, 7.26; S, 7.27. Found: C, 62.87; H, 4.22; Cl, 8.01; F, 4.39;
N, 6.56; O, 7.21; S, 7.17.
1. Woodward, R. B. In The Chemistry of Penicillin; Clarke, H. T., Johnson, J. R.,
Robinson, R., Eds.; Princeton University Press: Princeton, NJ, USA, 1949; p 443.
2. Crowfoot, D.; Bunn, C. W.; Rogers-Low, B. W.; Turner-Jones, A. In The Chemistry
of Penicillin; Clarke, H. T., Johnson, J. R., Robinson, R., Eds.; Princeton University
Press: Princeton, NJ, USA, 1949; pp 310–367.
5.2. Antimicrobial assays
3. Woodward, R. B. Science 1966, 153, 487.
4. Strominger, J. L. Antibiotics 1967, 1, 705.
5. Page, M. I. Acc. Chem. Res. 1984, 17, 144.
6. Mucsi, Z.; Chass, G. A.; Abranyi-Balogh, P.; Jozart, B.; Fang, D.-C.; Ramirez-
Cuesta, A. J.; Viskolcz, B.; Csizmadia, I. G. Phys. Chem. Chem. Phys. 2013, 15,
20447.
7. Neu, H. C. Science 1992, 257, 1064.
8. Cooper, M. A.; Shlaes, D. Nature 2011, 472, 32.
Antimicrobial activity for all isolates, with the exception of Mtb,
was evaluated by B. J. Plotkin and J. M. Green (Midwestern Univer-
sity, Downers Grove, IL). All organisms used are listed in Table 5
and were maintained at À80 °C. These bacteria, with the exception
of Mtb, were provided by J. Thjio (Loyola University Stritch School
of Medicine, Maywood, IL). The microdilution broth method was
used to determine the minimal inhibitory concentrations and the
minimum bactericidal concentrations (MIC/MBC).31 All lactams
were dissolved in dimethylsufamethoxizone (DMSO), then diluted
at least 10-fold into Mueller–Hinton (MH) broth. Each compound
9. Turos, E.; Konaklieva, M. I.; Ren, R.; Shi, H.; Gonzalez, J.; Dickey, S.; Lim, D. V.
Tetrahedron 2000, 56, 5571.
10. Turos, E.; Long, T. E.; Konaklieva, M. I.; Coates, C.; Shim, Y.-J.; Dickey, S.; Lim, D.
V.; Cannons, A. Bioorg. Med. Chem. Lett. 2002, 12, 2229.
11. Long, T. E.; Turos, E.; Konaklieva, M. I.; Blum, A. E.; Amry, A.; Baker, E. A.;
Suwandi, L. S.; McCain, M. D.; Rahman, M. F.; Dickey, S., et al. Bioorg. Med. Chem.
2003, 11, 1859.
12. Bhattacharya, B.; Turos, E. Tetrahedron 2012, 68, 10665.
13. Revell, K. D.; Heldreth, B.; Long, T. E.; Jang, S.; Turos, E. Bioorg. Med. Chem. 2007,
15, 2453.
14. Staub, I.; Sieber, S. A. J. Am. Chem. Soc. 2008, 130, 13400.
15. Staub, I.; Sieber, S. A. J. Am. Chem. Soc. 2009, 131, 6271.
16. Mehta, P. D.; Sengar, N. P. S.; Pathak, A. K. Eur. J. Med. Chem. 2010, 45, 5541.
17. Galletti, P.; Giacomini, D. Curr. Med. Chem. 2011, 18, 4265.
18. Toraskar, M.; Kulkarni, V.; Kadam, V. J. Pharm. Res. 2010, 3, 169.
19. Kostova, M. B.; Myers, C. J.; Beck, T. N.; Plotkin, B. J.; Green, J. M.; Boshoff, H. M.;
Barry, C., III; Deschamps, J.; Konaklieva, M. I. Bioorg. Med. Chem. 2011, 19, 6842.
20. Clauss, K.; Grimm, D.; Prossel, G. Justus Liebigs Ann. Chem. 1974, 4, 539.
21. Wasserman, H. H.; Xia, M.; Carr, A. J.; Han, W. T.; Siegel, M. G. Tetrahedron 2000,
56, 5621.
was then serially diluted in MH (100
tions, between 500 g/ml and 15.6
2-fold dilutions of DMSO alone. Bacteria were added to each well
(5–105 cells/ml; 100
L MH/well). After incubation (37 °C; 48 h)
lL/well; series of 2-fold dilu-
l
lg/ml). Controls consisted of
l
wells were scored for growth at 24 h and 48 h. The MIC was
defined as the lowest concentration of drug at which no growth
was observed. The MBC was determined as the lowest concentra-
tion of drug at which no growth as measured by standard plating
(10 lL, MH agar) of wells where no growth was observed. All tests
were performed in quadruplicate.
Mycobacterium tuberculosis (Mtb) H37Rv was used for all Mtb
susceptibility determinations. These determinations were done
by Kriti Arora (Tuberculosis Research Section, NIAID, NIH Bethesda,
MD).
22. Substituted azetidin-2-ones for treatment of atherosclerosis Patent:
US5990102 A1, 1999.
23. Mulchande, J.; Martins, L.; Moreira, R.; Archer, M.; Oliveira, T. F.; Iley, J. Org.
Biomol. Chem. 2007, 5, 2617.
ˇ
ˇ
24. Riley, K. E.; Murray, J. S.; Fanfrlík, J.; Rezác, J.; Solá, R. J.; Concha, M. C.; Ramos, F.
M.; Politzer, P. J. Mol. Model. 2013, 19, 4651.
Minimum inhibitory concentrations were done determined by
microdilution broth method as described previously.32
25. Su, Z.; Peng, L.; Worthington, R.; Melander, C. ChemMedChem 2011, 6, 2243.
26. Rogers, S., 3rd; Huigens, R. W.; Cavenagh, J.; Melander, C. Antimicrob. Agents
Chemother. 2010, 54, 2112.
Acknowledgments
27. Gu, H. Z.; Fedor, L. R. J. Org. Chem. 1990, 55, 5655.
28. The Organic Chemistry of b-Lactams; Georg, G. I., Ed.; VCH: New York, NY, USA,
1993.
29. Chemistry and Biology of b-Lactam Antibiotics; Morin, R. B., Gorman, M., Eds.;
Academic Press: New York, NY, USA, 1982; Vols. 1–3,.
30. Deziel, R.; Malenfant, E. Bioorg. Med. Chem. Lett. 1998, 8, 1437.
31. National Committee for Clinical Laboratory Standards, 1997.
32. Wong, S. Y.; Lee, J. S.; Kwak, H. K.; Via, L. E.; Boshoff, H. I.; Barry, C. E., III
Antimicrob. Agents Chemother. 2011, 55, 2515.
This work was funded in part, by the American University
Research Grant and the Department of Chemistry at American Uni-
versity, Washington, D.C. and by Midwestern University, Downers
Grove, IL. This research was supported in part by the Intramural
Research Program of the NIH, NIAID.